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Sample records for esterase e3 gene

  1. Cloning, Characterization, Controlled Overexpression, and Inactivation of the Major Tributyrin Esterase Gene of Lactococcus lactis

    NARCIS (Netherlands)

    Fernández, Leonides; Beerthuyzen, Marke M.; Brown, Julie; Siezen, Roland J.; Coolbear, Tim; Holland, Ross; Kuipers, Oscar P.

    2000-01-01

    The gene encoding the major intracellular tributyrin esterase of Lactococcus lactis was cloned using degenerate DNA probes based on 19 known N-terminal amino acid residues of the purified enzyme. The gene, named estA, was sequenced and found to encode a protein of 258 amino acid residues. The

  2. Enhanced biosurfactant production through cloning of three genes and role of esterase in biosurfactant release

    Science.gov (United States)

    2011-01-01

    Background Biosurfactants have been reported to utilize a number of immiscible substrates and thereby facilitate the biodegradation of panoply of polyaromatic hydrocarbons. Olive oil is one such carbon source which has been explored by many researchers. However, studying the concomitant production of biosurfactant and esterase enzyme in the presence of olive oil in the Bacillus species and its recombinants is a relatively novel approach. Results Bacillus species isolated from endosulfan sprayed cashew plantation soil was cultivated on a number of hydrophobic substrates. Olive oil was found to be the best inducer of biosurfactant activity. The protein associated with the release of the biosurfactant was found to be an esterase. There was a twofold increase in the biosurfactant and esterase activities after the successful cloning of the biosurfactant genes from Bacillus subtilis SK320 into E.coli. Multiple sequence alignment showed regions of similarity and conserved sequences between biosurfactant and esterase genes, further confirming the symbiotic correlation between the two. Biosurfactants produced by Bacillus subtilis SK320 and recombinant strains BioS a, BioS b, BioS c were found to be effective emulsifiers, reducing the surface tension of water from 72 dynes/cm to as low as 30.7 dynes/cm. Conclusion The attributes of enhanced biosurfactant and esterase production by hyper-producing recombinant strains have many utilities from industrial viewpoint. This study for the first time has shown a possible association between biosurfactant production and esterase activity in any Bacillus species. Biosurfactant-esterase complex has been found to have powerful emulsification properties, which shows promising bioremediation, hydrocarbon biodegradation and pharmaceutical applications. PMID:21707984

  3. Comparison of esterase gene amplification, gene expression and esterase activity in insecticide susceptible and resistant strains of the brown planthopper, Nilaparvata lugens (Stål).

    Science.gov (United States)

    Vontas, J G; Small, G J; Hemingway, J

    2000-12-01

    Organophosphorus and carbamate insecticide resistance in Nilaparvata lugens is based on amplification of a carboxylesterase gene, Nl-EST1. An identical gene occurs in susceptible insects. Quantitative real-time PCR was used to demonstrate that Nl-EST1 is amplified 3-7-fold in the genome of resistant compared to susceptible planthoppers. Expression levels were similar to amplification levels, with 1-15-fold more Nl-EST1 mRNA in individual insects and 5-11-fold more Nl-EST1 mRNA in mass whole body homogenates of resistant females compared to susceptibles. These values corresponded to an 8-10-fold increase in esterase activity in the head and thorax of individual resistant insects. Although amplification, expression and activity levels of Nl-EST1 in resistant N. lugens were similar, the correlation between esterase activity and Nl-EST1 mRNA levels in resistant individuals was not linear.

  4. Molecular population genetics of the β-esterase gene cluster of ...

    Indian Academy of Sciences (India)

    We suggest that the demographic history (bottleneck and admixture of genetically differentiated populations) is the major factor shaping the pattern of nucleotide polymorphism in the -esterase gene cluster. However there are some 'footprints' of directional and balancing selection shaping specific distribution of nucleotide ...

  5. Expanding the feruloyl esterase gene family of Aspergillus niger by characterization of a feruloyl esterase, FaeC.

    Science.gov (United States)

    Dilokpimol, Adiphol; Mäkelä, Miia R; Mansouri, Sadegh; Belova, Olga; Waterstraat, Martin; Bunzel, Mirko; de Vries, Ronald P; Hildén, Kristiina S

    2017-07-25

    A feruloyl esterase (FAE) from Aspergillus niger N402, FaeC was heterologously produced in Pichia pastoris X-33 in a yield of 10mg/L. FaeC was most active at pH 7.0 and 50°C, and showed broad substrate specificity and catalyzed the hydrolysis of methyl 3,4-dimethoxycinnamate, ethyl ferulate, methyl ferulate, methyl p-coumarate, ethyl coumarate, methyl sinapate, and methyl caffeate. The enzyme released both ferulic acid and p-coumaric acid from wheat arabinoxylan and sugar beet pectin (up to 3mg/g polysaccharide), and acted synergistically with a commercial xylanase increasing the release of ferulic acid up to six-fold. The expression of faeC increased over time in the presence of feruloylated polysaccharides. Cinnamic, syringic, caffeic, vanillic and ferulic acid induced the expression of faeC. Overall expression of faeC was very low in all tested conditions, compared to two other A. niger FAE encoding genes, faeA and faeB. Our data showed that the fae genes responded differently towards the feruloylated polysaccharides and tested monomeric phenolic compounds suggesting that the corresponding FAE isoenzymes may target different substrates in a complementary manner. This may increase the efficiency of the degradation of diverse plant biomass. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. aes, the gene encoding the esterase B in Escherichia coli, is a powerful phylogenetic marker of the species

    Directory of Open Access Journals (Sweden)

    Tuffery Pierre

    2009-12-01

    Full Text Available Abstract Background Previous studies have established a correlation between electrophoretic polymorphism of esterase B, and virulence and phylogeny of Escherichia coli. Strains belonging to the phylogenetic group B2 are more frequently implicated in extraintestinal infections and include esterase B2 variants, whereas phylogenetic groups A, B1 and D contain less virulent strains and include esterase B1 variants. We investigated esterase B as a marker of phylogeny and/or virulence, in a thorough analysis of the esterase B-encoding gene. Results We identified the gene encoding esterase B as the acetyl-esterase gene (aes using gene disruption. The analysis of aes nucleotide sequences in a panel of 78 reference strains, including the E. coli reference (ECOR strains, demonstrated that the gene is under purifying selection. The phylogenetic tree reconstructed from aes sequences showed a strong correlation with the species phylogenetic history, based on multi-locus sequence typing using six housekeeping genes. The unambiguous distinction between variants B1 and B2 by electrophoresis was consistent with Aes amino-acid sequence analysis and protein modelling, which showed that substituted amino acids in the two esterase B variants occurred mostly at different sites on the protein surface. Studies in an experimental mouse model of septicaemia using mutant strains did not reveal a direct link between aes and extraintestinal virulence. Moreover, we did not find any genes in the chromosomal region of aes to be associated with virulence. Conclusion Our findings suggest that aes does not play a direct role in the virulence of E. coli extraintestinal infection. However, this gene acts as a powerful marker of phylogeny, illustrating the extensive divergence of B2 phylogenetic group strains from the rest of the species.

  7. Genome-wide analysis of esterase-like genes in the striped rice stem borer, Chilo suppressalis.

    Science.gov (United States)

    Wang, Baoju; Wang, Ying; Zhang, Yang; Han, Ping; Li, Fei; Han, Zhaojun

    2015-06-01

    The striped rice stem borer, Chilo suppressalis, a destructive pest of rice, has developed high levels of resistance to certain insecticides. Esterases are reported to be involved in insecticide resistance in several insects. Therefore, this study systematically analyzed esterase-like genes in C. suppressalis. Fifty-one esterase-like genes were identified in the draft genomic sequences of the species, and 20 cDNA sequences were derived which encoded full- or nearly full-length proteins. The putative esterase proteins derived from these full-length genes are overall highly diversified. However, key residues that are functionally important including the serine residue in the active site are conserved in 18 out of the 20 proteins. Phylogenetic analysis revealed that most of these genes have homologues in other lepidoptera insects. Genes CsuEst6, CsuEst10, CsuEst11, and CsuEst51 were induced by the insecticide triazophos, and genes CsuEst9, CsuEst11, CsuEst14, and CsuEst51 were induced by the insecticide chlorantraniliprole. Our results provide a foundation for future studies of insecticide resistance in C. suppressalis and for comparative research with esterase genes from other insect species.

  8. Paralogous gene analysis reveals a highly enantioselective 1,2-O-isopropylideneglycerol caprylate esterase of Bacillus subtilis

    NARCIS (Netherlands)

    Droge, MJ; Bos, R; Quax, WJ

    Carboxylesterase NP of Bacillus subtilis Thai 1-8, characterized in 1992 as a very enantioselective (S)-naproxen esterase, was found to show no enantiopreference towards (S)-1,2-O-isopropylideneglycerol (IPG) esters. The ybfK gene was identified by the B. subtilis genome project as an unknown gene

  9. Cytological localization of adenosine kinase, nucleoside phosphorylase-1, and esterase-10 genes on mouse chromosome 14

    International Nuclear Information System (INIS)

    Samuelson, L.C.; Farber, R.A.

    1985-01-01

    The authors have determined the regional locations on mouse chromosome 14 of the genes for mouse adenosine kinase (ADK), nucleoside phosphorylase- 1 (NP-1), and esterase-10 (ES-10) by analysis of rearranged mouse chromosomes in gamma-irradiated Chinese hamster X mouse hybrid cell lines. Irradiated clones were screened for expression of the murine forms of these enzymes; segregant clones that expressed only one or two of the three markers were karyotyped. The patterns of enzyme expression in these segregants were correlated with the presence of rearranged chromosomes. The Adk gene was localized to bands A2 to B, Np-1 to bands B to C1, and Es-10 to bands D2 to E2

  10. A novel feruloyl esterase from rumen microbial metagenome: Gene cloning and enzyme characterization in the release of mono- and diferulic acids

    Science.gov (United States)

    A feruloyl esterase (FAE) gene was isolated from a rumen microbial metagenome, cloned into E. coli, and expressed in active form. The enzyme (RuFae4) was classified as a Type D feruloyl esterase based on its action on synthetic substrates and ability to release diferulates. The RuFae4 alone releas...

  11. Map-Based Cloning of the Gene Associated With the Soybean Maturity Locus E3

    Science.gov (United States)

    Watanabe, Satoshi; Hideshima, Rumiko; Xia, Zhengjun; Tsubokura, Yasutaka; Sato, Shusei; Nakamoto, Yumi; Yamanaka, Naoki; Takahashi, Ryoji; Ishimoto, Masao; Anai, Toyoaki; Tabata, Satoshi; Harada, Kyuya

    2009-01-01

    Photosensitivity plays an essential role in the response of plants to their changing environments throughout their life cycle. In soybean [Glycine max (L.) Merrill], several associations between photosensitivity and maturity loci are known, but only limited information at the molecular level is available. The FT3 locus is one of the quantitative trait loci (QTL) for flowering time that corresponds to the maturity locus E3. To identify the gene responsible for this QTL, a map-based cloning strategy was undertaken. One phytochrome A gene (GmPhyA3) was considered a strong candidate for the FT3 locus. Allelism tests and gene sequence comparisons showed that alleles of Misuzudaizu (FT3/FT3; JP28856) and Harosoy (E3/E3; PI548573) were identical. The GmPhyA3 alleles of Moshidou Gong 503 (ft3/ft3; JP27603) and L62-667 (e3/e3; PI547716) showed weak or complete loss of function, respectively. High red/far-red (R/FR) long-day conditions enhanced the effects of the E3/FT3 alleles in various genetic backgrounds. Moreover, a mutant line harboring the nonfunctional GmPhyA3 flowered earlier than the original Bay (E3/E3; PI553043) under similar conditions. These results suggest that the variation in phytochrome A may contribute to the complex systems of soybean flowering response and geographic adaptation. PMID:19474204

  12. Cloning and characterization of a pyrethroid pesticide decomposing esterase gene, Est3385, from Rhodopseudomonas palustris PSB-S.

    Science.gov (United States)

    Luo, Xiangwen; Zhang, Deyong; Zhou, Xuguo; Du, Jiao; Zhang, Songbai; Liu, Yong

    2018-05-09

    Full length open reading frame of pyrethroid detoxification gene, Est3385, contains 963 nucleotides. This gene was identified and cloned based on the genome sequence of Rhodopseudomonas palustris PSB-S available at the GneBank. The predicted amino acid sequence of Est3385 shared moderate identities (30-46%) with the known homologous esterases. Phylogenetic analysis revealed that Est3385 was a member in the esterase family I. Recombinant Est3385 was heterologous expressed in E. coli, purified and characterized for its substrate specificity, kinetics and stability under various conditions. The optimal temperature and pH for Est3385 were 35 °C and 6.0, respectively. This enzyme could detoxify various pyrethroid pesticides and degrade the optimal substrate fenpropathrin with a Km and Vmax value of 0.734 ± 0.013 mmol·l -1 and 0.918 ± 0.025 U·µg -1 , respectively. No cofactor was found to affect Est3385 activity but substantial reduction of enzymatic activity was observed when metal ions were applied. Taken together, a new pyrethroid degradation esterase was identified and characterized. Modification of Est3385 with protein engineering toolsets should enhance its potential for field application to reduce the pesticide residue from agroecosystems.

  13. A novel NR2E3 gene mutation in autosomal recessive retinitis pigmentosa with cystic maculopathy

    OpenAIRE

    Mahajan, D.; Votruba, Marcela

    2017-01-01

    NR2E3 is a gene that encodes for photoreceptor cell specific nuclear receptor, which is involved in cone proliferation. The splice site mutation 119-2A>C in NR2E3 (15q23) has been previously reported to underlie recessive enhanced cone S sensitivity syndrome, clumped pigmentary retinal degeneration, Goldman-Favre syndrome and also autosomal dominant and autosomal recessive retinitis pigmentosa (RP). However, the mutation c 571 + 2 T > C in NR2E3 has not been previously reported with retinal d...

  14. The Orf virus E3L homologue is able to complement deletion of the vaccinia virus E3L gene in vitro but not in vivo

    International Nuclear Information System (INIS)

    Vijaysri, Sangeetha; Talasela, Latha; Mercer, Andrew A.; Mcinnes, Colin J.; Jacobs, Bertram L.; Langland, Jeffrey O.

    2003-01-01

    Orf virus (OV), the prototypic parapoxvirus, is resistant to the effects of interferon (IFN) and this function of OV has been mapped to the OV20.0L gene. The protein product of this gene shares 31% amino acid identity to the E3L-encoded protein of vaccinia virus (VV) that is required for the broad host range and IFN-resistant phenotype of VV in cells in culture and for virulence of the virus in vivo. In this study we investigated whether the distantly related OV E3L homologue could complement the deletion of E3L in VV. The recombinant VV (VV/ORF-E3L) expressing the OV E3L homologue in place of VV E3L was indistinguishable from wt VV in its cell-culture phenotype. But VV/ORF-E3L was over a 1000-fold less pathogenic than wt VV (LD 50 > 5 x 10 6 PFU, compared to LD 50 of wtVV = 4 x 10 3 PFU) following intranasal infection of mice. While wt VV spread to the lungs and brain and replicated to high titers in the brain of infected mice, VV/ORF-E3L could not be detected in the lungs or brain following intranasal infection. VV/ORF-E3L was at least 100,000-fold less pathogenic than wt VV on intracranial injection. Domain swap experiments demonstrate that the difference in pathogenesis maps to the C-terminal domain of these proteins. This domain has been shown to be required for the dsRNA binding function of the VV E3L

  15. Towards the industrialization of new biosurfactants: Biotechnological opportunities for the lactone esterase gene from Starmerella bombicola.

    Science.gov (United States)

    Roelants, Sophie L K W; Ciesielska, Katarzyna; De Maeseneire, Sofie L; Moens, Helena; Everaert, Bernd; Verweire, Stijn; Denon, Quenten; Vanlerberghe, Brecht; Van Bogaert, Inge N A; Van der Meeren, Paul; Devreese, Bart; Soetaert, Wim

    2016-03-01

    Although sophorolipids (SLs) produced by S. bombicola are a real showcase for the industrialization of microbial biosurfactants, some important drawbacks are associated with this efficient biological process, e.g., the simultaneous production of acidic and lactonic SLs. Depending on the application, there is a requirement for the naturally produced mixture to be manipulated to give defined ratios of the components. Recently, the enzyme responsible for the lactonization of SLs was discovered. The discovery of the gene encoding this lactone esterase (sble) enabled the development of promising S. bombicola strains producing either solely lactonic (using a sble overexpression strain described in this paper: oe sble) or solely acidic SLs (using a sble deletion strain, which was recently described, but not characterized yet: Δsble). The new S. bombicola strains were used to investigate the production processes (fermentation and purification) of either lactonic or acidic SLs. The strains maintain the high inherent productivities of the wild-type or even perform slightly better and thus represent a realistic industrial opportunity. 100% acidic SLs with a mixed acetylation pattern were obtained for the Δsble strain, while the inherent capacity to selectively produce lactonic SLs was significantly increased (+42%) for the oe sble strain (99% lactonic SLs). Moreover, the regulatory effect of citrate on lactone SL formation for the wild-type was absent in this new strain, which indicates that it is more robust and better suited for the industrial production of lactonic SLs. Basic parameters were determined for the purified SLs, which confirm that the two new strains produce molecules with distinctive properties of which the application potential can now easily be investigated independently. © 2015 Wiley Periodicals, Inc.

  16. A novel esterase from Saccharomyces carlsbergensis, a possible function for the yeast TIP1 gene

    DEFF Research Database (Denmark)

    Horsted, M W; Dey, E S; Holmberg, S

    1998-01-01

    An extracellular esterase was isolated from the brewer's yeast, Saccharomyces carlsbergensis. Inhibition by diisopropyl fluorophosphate shows that the enzyme has a serine active site. By mass spectrometry, the molecular weight of the enzyme was 16.9 kDa. The optimal pH for activity was in the range...

  17. The Aspergillus niger faeB gene encodes a second feruloyl esterase involved in pectin and xylan degradation and is specifically induced in the presence of aromatic compounds

    NARCIS (Netherlands)

    Vries, de R.P.; vanKuyk, P.A.; Kester, H.C.M.; Visser, J.

    2002-01-01

    The faeB gene encoding a second feruloyl esterase from Aspergillus niger has been cloned and characterized. It consists of an open reading frame of 1644 bp containing one intron. The gene encodes a protein of 521 amino acids that has sequence similarity to that of an Aspergillus oryzae tannase.

  18. Intron retention regulates the expression of pectin methyl esterase inhibitor (Pmei) genes during wheat growth and development.

    Science.gov (United States)

    Rocchi, V; Janni, M; Bellincampi, D; Giardina, T; D'Ovidio, R

    2012-03-01

    Pectin is an important component of the plant cell wall and its remodelling occurs during normal plant growth or following stress responses. Pectin is secreted into the cell wall in a highly methyl-esterified form and subsequently de-methyl-esterified by pectin methyl esterase (PME), whose activity is controlled by the pectin methyl esterase inhibitor protein (PMEI). Cereal cell wall contains a low amount of pectin; nonetheless the level and pattern of pectin methyl esterification play a primary role during development or pathogen infection. Since few data are available on the role of PMEI in plant development and defence of cereal species, we isolated and characterised three Pmei genes (Tdpmei2.1, Tdpmei2.2 and Tdpmei3) and their encoded products in wheat. Sequence comparisons showed a low level of intra- and inter-specific sequence conservation of PMEIs. Tdpmei2.1 and Tdpmei2.2 share 94% identity at protein level, but only 20% identity with the product of Tdpmei3. All three Tdpmei genes code for functional inhibitors of plant PMEs and do not inhibit microbial PMEs or a plant invertase. RT-PCR analyses demonstrated, for the first time to our knowledge, that Pmei genes are regulated by intron retention. Processed and unprocessed transcripts of Tdpmei2.1 and Tdpmei2.2 accumulated in several organs, but anthers contained only mature transcripts. Tdpmei3 lacks introns and its transcript accumulated mainly in stem internodes. These findings suggest that products encoded by these Tdpmei genes control organ- or tissue-specific activity of specific PME isoforms in wheat. © 2011 German Botanical Society and The Royal Botanical Society of the Netherlands.

  19. Identification and characterization of a GDSL esterase gene located proximal to the swr quorum-sensing system of Serratia liquefaciens MG1

    DEFF Research Database (Denmark)

    Riedel, K.; Talker-Huiber, D.; Givskov, Michael Christian

    2003-01-01

    direction, designated estA, which encodes an esterase that belongs to family II of lipolytic enzymes. EstA was heterologously expressed in Escherichia coli, and the substrate specificity of the enzyme was determined in crude extracts. With the aid of zymograms visualizing EstA on polyacrylamide gels...... and by the analysis of a transcriptional fusion of the estA promoter to the promoterless lux4B genes, we showed that expression of the esterase is not regulated by the swr quorum-sensing system. An estA mutant was generated and was found to exhibit growth defects on minimal medium containing Tween 20 or Tween 80...

  20. Esterase Isoenzyme Variants in Barley

    DEFF Research Database (Denmark)

    Hvid, S.; Nielsen, G.

    1977-01-01

    Gene symbols are proposed for 27 esterase isoenzyme alleles representing 10 loci in barley. Two new esterase loci, Est 9 and Est 10, each with an active and a silent allele, and three new alleles in previously described loci were found. A few chemical and physical characteristics of the different...... esterase isoenzyme systems were studied. The heat inactivation temperature differed for the isoenzymes coded by most of the loci, whereas the substrate and inhibitor specificity of the isoenzymes was less distinct. A possible relationship between some of the systems is discussed....

  1. Multifunctionality and diversity of GDSL esterase/lipase gene family in rice (Oryza sativa L. japonica genome: new insights from bioinformatics analysis

    Directory of Open Access Journals (Sweden)

    Chepyshko Hanna

    2012-07-01

    Full Text Available Abstract Background GDSL esterases/lipases are a newly discovered subclass of lipolytic enzymes that are very important and attractive research subjects because of their multifunctional properties, such as broad substrate specificity and regiospecificity. Compared with the current knowledge regarding these enzymes in bacteria, our understanding of the plant GDSL enzymes is very limited, although the GDSL gene family in plant species include numerous members in many fully sequenced plant genomes. Only two genes from a large rice GDSL esterase/lipase gene family were previously characterised, and the majority of the members remain unknown. In the present study, we describe the rice OsGELP (Oryza sativa GDSL esterase/lipase protein gene family at the genomic and proteomic levels, and use this knowledge to provide insights into the multifunctionality of the rice OsGELP enzymes. Results In this study, an extensive bioinformatics analysis identified 114 genes in the rice OsGELP gene family. A complete overview of this family in rice is presented, including the chromosome locations, gene structures, phylogeny, and protein motifs. Among the OsGELPs and the plant GDSL esterase/lipase proteins of known functions, 41 motifs were found that represent the core secondary structure elements or appear specifically in different phylogenetic subclades. The specification and distribution of identified putative conserved clade-common and -specific peptide motifs, and their location on the predicted protein three dimensional structure may possibly signify their functional roles. Potentially important regions for substrate specificity are highlighted, in accordance with protein three-dimensional model and location of the phylogenetic specific conserved motifs. The differential expression of some representative genes were confirmed by quantitative real-time PCR. The phylogenetic analysis, together with protein motif architectures, and the expression profiling were

  2. Functional Cloning and Expression of the Schizophyllum commune Glucuronoyl Esterase Gene and Characterization of the Recombinant Enzyme

    Science.gov (United States)

    Wong, Dominic W. S.; Chan, Victor J.; McCormack, Amanda A.; Hirsch, Ján; Biely, Peter

    2012-01-01

    The gene encoding Schizophyllum commune glucuronoyl esterase was identified in the scaffold 17 of the genome, containing two introns of 50 bp and 48 bp, with a transcript sequence of 1179 bp. The gene was synthesized and cloned into Pichia pastoris expression vector pGAPZα to achieve constitutive expression and secretion of the recombinant enzyme in soluble active form. The purified protein was 53 kD with glycosylation and had an acidic pI of 3.7. Activity analysis on several uronic acids and their derivatives suggests that the enzyme recognized only esters of 4-O-methyl-D-glucuronic acid derivatives, even with a 4-nitrophenyl aglycon but did not hydrolyze the ester of D-galacturonic acid. The kinetic values were K m 0.25 mM, V max 16.3 μM·min−1, and k cat 9.27 s−1 with 4-nitrophenyl 2-O-(methyl 4-O-methyl-α-D-glucopyranosyluronate)-β-D-xylopyranoside as the substrate. PMID:22844600

  3. The Aspergillus niger faeB gene encodes a second feruloyl esterase involved in pectin and xylan degradation and is specifically induced in the presence of aromatic compounds.

    Science.gov (United States)

    de Vries, Ronald P; vanKuyk, Patricia A; Kester, Harry C M; Visser, Jaap

    2002-04-15

    The faeB gene encoding a second feruloyl esterase from Aspergillus niger has been cloned and characterized. It consists of an open reading frame of 1644 bp containing one intron. The gene encodes a protein of 521 amino acids that has sequence similarity to that of an Aspergillus oryzae tannase. However, the encoded enzyme, feruloyl esterase B (FAEB), does not have tannase activity. Comparison of the physical characteristics and substrate specificity of FAEB with those of a cinnamoyl esterase from A. niger [Kroon, Faulds and Williamson (1996) Biotechnol. Appl. Biochem. 23, 255-262] suggests that they are in fact the same enzyme. The expression of faeB is specifically induced in the presence of certain aromatic compounds, but not in the presence of other constituents present in plant-cell-wall polysaccharides such as arabinoxylan or pectin. The expression profile of faeB in the presence of aromatic compounds was compared with the expression of A. niger faeA, encoding feruloyl esterase A (FAEA), and A. niger bphA, the gene encoding a benzoate-p-hydroxylase. All three genes have different subsets of aromatic compounds that induce their expression, indicating the presence of different transcription activating systems in A. niger that respond to aromatic compounds. Comparison of the activity of FAEA and FAEB on sugar-beet pectin and wheat arabinoxylan demonstrated that they are both involved in the degradation of both polysaccharides, but have opposite preferences for these substrates. FAEA is more active than FAEB towards wheat arabinoxylan, whereas FAEB is more active than FAEA towards sugar-beet pectin.

  4. Molecular Cloning and Characterization of a Newly Isolated Pyrethroid-Degrading Esterase Gene from a Genomic Library of Ochrobactrum anthropi YZ-1

    Science.gov (United States)

    Song, Jinlong; Shi, Yanhua; Li, Kang; Zhao, Bin; Yan, Yanchun

    2013-01-01

    A novel pyrethroid-degrading esterase gene pytY was isolated from the genomic library of Ochrobactrum anthropi YZ-1. It possesses an open reading frame (ORF) of 897 bp. Blast search showed that its deduced amino acid sequence shares moderate identities (30% to 46%) with most homologous esterases. Phylogenetic analysis revealed that PytY is a member of the esterase VI family. pytY showed very low sequence similarity compared with reported pyrethroid-degrading genes. PytY was expressed, purified, and characterized. Enzyme assay revealed that PytY is a broad-spectrum degrading enzyme that can degrade various pyrethroids. It is a new pyrethroid-degrading gene and enriches genetic resource. Kinetic constants of Km and Vmax were 2.34 mmol·L−1 and 56.33 nmol min−1, respectively, with lambda-cyhalothrin as substrate. PytY displayed good degrading ability and stability over a broad range of temperature and pH. The optimal temperature and pH were of 35°C and 7.5. No cofactors were required for enzyme activity. The results highlighted the potential use of PytY in the elimination of pyrethroid residuals from contaminated environments. PMID:24155944

  5. Insertion of an esterase gene into a specific locust pathogen (Metarhizium acridum enables it to infect caterpillars.

    Directory of Open Access Journals (Sweden)

    Sibao Wang

    2011-06-01

    Full Text Available An enduring theme in pathogenic microbiology is poor understanding of the mechanisms of host specificity. Metarhizium is a cosmopolitan genus of invertebrate pathogens that contains generalist species with broad host ranges such as M. robertsii (formerly known as M. anisopliae var. anisopliae as well as specialists such as the acridid-specific grasshopper pathogen M. acridum. During growth on caterpillar (Manduca sexta cuticle, M. robertsii up-regulates a gene (Mest1 that is absent in M. acridum and most other fungi. Disrupting M. robertsii Mest1 reduced virulence and overexpression increased virulence to caterpillars (Galleria mellonella and M. sexta, while virulence to grasshoppers (Melanoplus femurrubrum was unaffected. When Mest1 was transferred to M. acridum under control of its native M. robertsii promoter, the transformants killed and colonized caterpillars in a similar fashion to M. robertsii. MEST1 localized exclusively to lipid droplets in M. robertsii conidia and infection structures was up-regulated during nutrient deprivation and had esterase activity against lipids with short chain fatty acids. The mobilization of stored lipids was delayed in the Mest1 disruptant mutant. Overall, our results suggest that expression of Mest1 allows rapid hydrolysis of stored lipids, and promotes germination and infection structure formation by M. robertsii during nutrient deprivation and invasion, while Mest1 expression in M. acridum broadens its host range by bypassing the regulatory signals found on natural hosts that trigger the mobilization of endogenous nutrient reserves. This study suggests that speciation in an insect pathogen could potentially be driven by host shifts resulting from changes in a single gene.

  6. Involvement of Three Esterase Genes from Panonychus citri (McGregor in Fenpropathrin Resistance

    Directory of Open Access Journals (Sweden)

    Xiao-Min Shen

    2016-08-01

    Full Text Available The citrus red mite, Panonychus citri (McGregor, is a major citrus pest with a worldwide distribution and an extensive record of pesticide resistance. However, the underlying molecular mechanism associated with fenpropathrin resistance in this species have not yet been reported. In this study, synergist triphenyl phosphate (TPP dramatically increased the toxicity of fenpropathrin, suggesting involvement of carboxylesterases (CarEs in the metabolic detoxification of this insecticide. The subsequent spatiotemporal expression pattern analysis of PcE1, PcE7 and PcE9 showed that three CarEs genes were all over-expressed after insecticide exposure and higher transcripts levels were observed in different field resistant strains of P. citri. Heterologous expression combined with 3-(4,5-dimethyl-thiazol-2-yl-2,5-diphenyltetra-zolium bromide (MTT cytotoxicity assay in Spodoptera frugiperda (Sf9 cells revealed that PcE1-, PcE7- or PcE9-expressing cells showed significantly higher cytoprotective capability than parental Sf9 cells against fenpropathrin, demonstrating that PcEs probably detoxify fenpropathrin. Moreover, gene silencing through the method of leaf-mediated dsRNA feeding followed by insecticide bioassay increased the mortalities of fenpropathrin-treated mites by 31% (PcE1, 27% (PcE7 and 22% (PcE9, respectively, after individual PcE gene dsRNA treatment. In conclusion, this study provides evidence that PcE1, PcE7 and PcE9 are functional genes mediated in fenpropathrin resistance in P. citri and enrich molecular understanding of CarEs during the resistance development of the mite.

  7. Genome-wide characterization of pectin methyl esterase genes reveals members differentially expressed in tolerant and susceptible wheats in response to Fusarium graminearum.

    Science.gov (United States)

    Zega, Alessandra; D'Ovidio, Renato

    2016-11-01

    Pectin methyl esterase (PME) genes code for enzymes that are involved in structural modifications of the plant cell wall during plant growth and development. They are also involved in plant-pathogen interaction. PME genes belong to a multigene family and in this study we report the first comprehensive analysis of the PME gene family in bread wheat (Triticum aestivum L.). Like in other species, the members of the TaPME family are dispersed throughout the genome and their encoded products retain the typical structural features of PMEs. qRT-PCR analysis showed variation in the expression pattern of TaPME genes in different tissues and revealed that these genes are mainly expressed in flowering spikes. In our attempt to identify putative TaPME genes involved in wheat defense, we revealed a strong variation in the expression of the TaPME following Fusarium graminearum infection, the causal agent of Fusarium head blight (FHB). Particularly interesting was the finding that the expression profile of some PME genes was markedly different between the FHB-resistant wheat cultivar Sumai3 and the FHB-susceptible cultivar Bobwhite, suggesting a possible involvement of these PME genes in FHB resistance. Moreover, the expression analysis of the TaPME genes during F. graminearum progression within the spike revealed those genes that responded more promptly to pathogen invasion. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  8. A cold-induced pectin methyl-esterase inhibitor gene contributes negatively to freezing tolerance but positively to salt tolerance in Arabidopsis.

    Science.gov (United States)

    Chen, Jian; Chen, Xuehui; Zhang, Qingfeng; Zhang, Yidan; Ou, Xiangli; An, Lizhe; Feng, Huyuan; Zhao, Zhiguang

    2018-03-01

    Plant pectin methyl-esterase (PME) and PME inhibitor (PMEI) belong to large gene families whose members are proposed to be widely involved in growth, development, and stress responses; however, the biological functions of most PMEs and PMEIs have not been characterized. In this study, we studied the roles of CbPMEI1, a cold-induced pectin methyl-esterase inhibitor (PMEI) gene from Chorispora bungeana, under freezing and salt stress. The putative CbPMEI1 peptide shares highest similarity (83%) with AT5G62360 (PMEI13) of Arabidopsis. Overexpression of either CbPMEI1 or PMEI13 in Arabidopsis decreased tissue PME activity and enhanced the degree of methoxylation of cell wall pectins, indicating that both genes encode functional PMEIs. CbPMEI1 and PMEI13 were induced by cold but repressed by salt stress and abscisic acid, suggesting distinct roles of the genes in freezing and salt stress tolerance. Interestingly, transgenic Arabidopsis plants overexpressing CbPMEI1 or PMEI13 showed decreased freezing tolerance, as indicated by survival and electrolyte leakage assays. On the other hand, the salt tolerance of transgenic plants was increased, showing higher rates of germination, root growth, and survival under salinity conditions as compared with non-transgenic wild-type plants. Although the transgenic plants were freezing-sensitive, they showed longer roots than wild-type plants under cold conditions, suggesting a role of PMEs in balancing the trade-off between freezing tolerance and growth. Thus, our study indicates that CbPMEI1 and PMEI13 are involved in root growth regulation under cold and salt stresses, and suggests that PMEIs may be potential targets for genetic engineering aimed to improve fitness of plants under stress conditions. Copyright © 2018 Elsevier GmbH. All rights reserved.

  9. New truncation mutation of the NR2E3 gene in a Japanese patient with enhanced S-cone syndrome.

    Science.gov (United States)

    Kuniyoshi, Kazuki; Hayashi, Takaaki; Sakuramoto, Hiroyuki; Mishima, Hiroshi; Tsuneoka, Hiroshi; Tsunoda, Kazushige; Iwata, Takeshi; Shimomura, Yoshikazu

    2016-11-01

    The enhanced S-cone syndrome (ESCS) is a rare hereditary retinal degeneration that has enhanced short wavelength-sensitive cone (S-cone) functions. The longitudinal clinical course of this disease has been rarely reported, and the genetic aspects of ESCS have not been well investigated in the Japanese population. In this report, we present our clinical and genetic findings for 2 patients with ESCS. The patients were 2 unrelated Japanese men. Standard ophthalmic examinations and mutation screening for the NR2E3 gene were performed. Patient 1 was a 36-year-old man, and his clinical findings were typical of ESCS. His decimal best-corrected visual acuity (BCVA) was 1.0 OD and 0.5 OS after removal of cataracts. Genetic investigations revealed a homozygous truncation frameshift, the p.I307LfsX33 mutation. Patient 2 was an 11-year-old boy when he was first examined by us. His clinical findings were typical of ESCS except for uveitis in the left eye. His decimal BCVA at the age of 39 years was maintained at 1.5 in each eye, although the retinal degeneration and visual field impairments had progressed during the follow-up period. The genetic investigations revealed homozygous mutations of p.R104Q in the NR2E3 gene. The frameshift mutation, p.I307LfsX33, in the NR2E3 gene is a new causative mutation for ESCS. The clinical observations for patient 2 are the longest ever reported. The retinal degeneration caused by this mutation is slowly progressive, and these patients maintained good vision with maintenance of the foveal structure until their late thirties.

  10. A novel esterase gene cloned from a metagenomic library from neritic sediments of the South China Sea

    Science.gov (United States)

    2011-01-01

    Background Marine microbes are a large and diverse group, which are exposed to a wide variety of pressure, temperature, salinity, nutrient availability and other environmental conditions. They provide a huge potential source of novel enzymes with unique properties that may be useful in industry and biotechnology. To explore the lipolytic genetic resources in the South China Sea, 23 sediment samples were collected in the depth South China Sea sediments assemblage in plasmid vector containing about 194 Mb of community DNA was prepared. Screening of a part of the unamplified library resulted in isolation of 15 unique lipolytic clones with the ability to hydrolyze tributyrin. A positive recombinant clone (pNLE1), containing a novel esterase (Est_p1), was successfully expressed in E. coli and purified. In a series of assays, Est_p1 displayed maximal activity at pH 8.57, 40°C, with ρ-Nitrophenyl butyrate (C4) as substrate. Compared to other metagenomic esterases, Est_p1 played a notable role in specificity for substrate C4 (kcat/Km value 11,500 S-1m M-1) and showed no inhibited by phenylmethylsulfonyl fluoride, suggested that the substrate binding pocket was suitable for substrate C4 and the serine active-site residue was buried at the bottom of substrate binding pocket which sheltered by a lid structure. Conclusions Esterase, which specificity towards short chain fatty acids, especially butanoic acid, is commercially available as potent flavoring tools. According the outstanding activity and specificity for substrate C4, Est_p1 has potential application in flavor industries requiring hydrolysis of short chain esters. PMID:22067554

  11. Wounding coordinately induces cell wall protein, cell cycle and pectin methyl esterase genes involved in tuber closing layer and wound periderm development.

    Science.gov (United States)

    Neubauer, Jonathan D; Lulai, Edward C; Thompson, Asunta L; Suttle, Jeffrey C; Bolton, Melvin D

    2012-04-15

    Little is known about the coordinate induction of genes that may be involved in agriculturally important wound-healing events. In this study, wound-healing events were determined together with wound-induced expression profiles of selected cell cycle, cell wall protein, and pectin methyl esterase genes using two diverse potato genotypes and two harvests (NDTX4271-5R and Russet Burbank tubers; 2008 and 2009 harvests). By 5 d after wounding, the closing layer and a nascent phellogen had formed. Phellogen cell divisions generated phellem layers until cessation of cell division at 28 d after wounding for both genotypes and harvests. Cell cycle genes encoding epidermal growth factor binding protein (StEBP), cyclin-dependent kinase B (StCDKB) and cyclin-dependent kinase regulatory subunit (StCKS1At) were induced by 1 d after wounding; these expressions coordinated with related phellogen formation and the induction and cessation of phellem cell formation. Genes encoding the structural cell wall proteins extensin (StExt1) and extensin-like (StExtlk) were dramatically up-regulated by 1-5 d after wounding, suggesting involvement with closing layer and later phellem cell layer formation. Wounding up-regulated pectin methyl esterase genes (StPME and StPrePME); StPME expression increased during closing layer and phellem cell formation, whereas maximum expression of StPrePME occurred at 5-14 d after wounding, implicating involvement in later modifications for closing layer and phellem cell formation. The coordinate induction and expression profile of StTLRP, a gene encoding a cell wall strengthening "tyrosine-and lysine-rich protein," suggested a role in the formation of the closing layer followed by phellem cell generation and maturation. Collectively, the genes monitored were wound-inducible and their expression profiles markedly coordinated with closing layer formation and the index for phellogen layer meristematic activity during wound periderm development; results were more

  12. Glucuronoyl esterase--novel carbohydrate esterase produced by Schizophyllum commune.

    Science.gov (United States)

    Spániková, Silvia; Biely, Peter

    2006-08-21

    The cellulolytic system of the wood-rotting fungus Schizophyllum commune contains an esterase that hydrolyzes methyl ester of 4-O-methyl-d-glucuronic acid. The enzyme, called glucuronoyl esterase, was purified to electrophoretic homogeneity from a cellulose-spent culture fluid. Its substrate specificity was examined on a number of substrates of other carbohydrate esterases such as acetylxylan esterase, feruloyl esterase and pectin methylesterase. The glucuronoyl esterase attacks exclusively the esters of MeGlcA. The methyl ester of free or glycosidically linked MeGlcA was not hydrolysed by other carbohydrate esterases. The results suggest that we have discovered a new type of carbohydrate esterase that might be involved in disruption of ester linkages connecting hemicellulose and lignin in plant cell walls.

  13. BIIDXI, the At4g32460 DUF642 gene, is involved in pectin methyl esterase regulation during Arabidopsis thaliana seed germination and plant development.

    Science.gov (United States)

    Zúñiga-Sánchez, Esther; Soriano, Diana; Martínez-Barajas, Eleazar; Orozco-Segovia, Alma; Gamboa-deBuen, Alicia

    2014-12-02

    DUF642 proteins constitute a highly conserved family of proteins that are associated with the cell wall and are specific to spermatophytes. Transcriptome studies have suggested that members of this family are involved in seed development and germination processes. Previous in vitro studies have revealed that At4g32460- and At5g11420-encoded proteins interact with the catalytic domain of pectin methyl esterase 3 (AtPME3, which is encoded by At3g14310). PMEs play an important role in plant development, including seed germination. The aim of this study was to evaluate the function of the DUF642 gene At4g32460 during seed germination and plant development and to determine its relation to PME activity regulation. Our results indicated that the DUF642 proteins encoded by At4g32460 and At5g11420 could be positive regulators of PME activity during several developmental processes. Transgenic lines overexpressing these proteins showed increased PME activity during seed germination, and improved seed germination performance. In plants expressing At4g32460 antisense RNA, PME activity was decreased in the leaves, and the siliques were very short and contained no seeds. This phenotype was also present in the SALK_142260 and SALK_054867 lines for At4g32460. Our results suggested that the DUF642 family contributes to the complexity of the methylesterification process by participating in the fine regulation of pectin status during plant development.

  14. A whole genome screening and RNA interference identify a juvenile hormone esterase-like gene of the diamondback moth, Plutella xylostella.

    Science.gov (United States)

    Gu, Xiaojun; Kumar, Sunil; Kim, Eunjin; Kim, Yonggyun

    2015-09-01

    Juvenile hormone (JH) plays a crucial role in preventing precocious metamorphosis and stimulating reproduction. Thus, its hemolymph titer should be under a tight control. As a negative controller, juvenile hormone esterase (JHE) performs a rapid breakdown of residual JH in the hemolymph during last instar to induce a larval-to-pupal metamorphosis. A whole genome of the diamondback moth (DBM), Plutella xylostella, has been annotated and proposed 11 JHE candidates. Sequence analysis using conserved motifs commonly found in other JHEs proposed a putative JHE (Px004817). Px004817 (64.61 kDa, pI=5.28) exhibited a characteristic JHE expression pattern by showing high peak at the early last instar, at which JHE enzyme activity was also at a maximal level. RNA interference of Px004817 reduced JHE activity and interrupted pupal development with a significant increase of larval period. This study identifies Px004817 as a JHE-like gene of P. xylostella. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Novel recombinant ethyl ferulate esterase from Burkholderia multivorans

    CSIR Research Space (South Africa)

    Rashamuse, KJ

    2007-11-01

    Full Text Available Isolation and identification of bacterial isolates with specific ferulic acid (FA) esterase activity and cloning of a gene encoding activity. A micro-organism with ethyl ferulate hydrolysing (EFH) activity was isolated by culture enrichment...

  16. E3B1, a human homologue of the mouse gene product Abi-1, sensitizes activation of Rap1 in response to epidermal growth factor

    International Nuclear Information System (INIS)

    Jenei, Veronika; Andersson, Tommy; Jakus, Judit; Dib, Karim

    2005-01-01

    E3B1, a human homologue of the mouse gene product Abi-1, has been implicated in growth-factor-mediated regulation of the small GTPases p21 Ras and Rac. E3b1 is a regulator of Rac because it can form a complex with Sos-1 and eps8, and such a Sos-1-e3B1-eps8 complex serves as a guanine nucleotide exchange factor for Rac. In the present study, we found that overexpression of e3B1 in NIH3T3/EGFR cells sensitized EGF-induced activation of Rac1, whereas it had no impact on EGF-induced activation of p21 Ras . Remarkably, we found that EGF-induced activation of the p21 Ras -related GTPase Rap1 was also sensitized in NIH3T3/EGFR-e3B1 cells. Thus, in NIH3T3/EGFR-e3B1 cells, maximal EGF-induced activation of Rap1 occurs with a dose of EGF much lower than in NIH3T3/EGFR cells. We also report that overexpression of e3B1 in NIH3T3/EGFR cells renders EGF-induced activation of Rap1 completely dependent on Src tyrosine kinases but not on c-Abl. However, EGF-induced tyrosine phosphorylation of the Rap GEF C3G occurred regardless of whether e3B1 was overexpressed or not, and this did not involve Src tyrosine kinases. Accordingly, we propose that overexpression of e3B1 in NIH3T3/EGFR cells leads to mobilization of Src tyrosine kinases that participate in EGF-induced activation of Rap1 and inhibition of cell proliferation

  17. E3L and F1L Gene Functions Modulate the Protective Capacity of Modified Vaccinia Virus Ankara Immunization in Murine Model of Human Smallpox

    Directory of Open Access Journals (Sweden)

    Asisa Volz

    2018-01-01

    Full Text Available The highly attenuated Modified Vaccinia virus Ankara (MVA lacks most of the known vaccinia virus (VACV virulence and immune evasion genes. Today MVA can serve as a safety-tested next-generation smallpox vaccine. Yet, we still need to learn about regulatory gene functions preserved in the MVA genome, such as the apoptosis inhibitor genes F1L and E3L. Here, we tested MVA vaccine preparations on the basis of the deletion mutant viruses MVA-ΔF1L and MVA-ΔE3L for efficacy against ectromelia virus (ECTV challenge infections in mice. In non-permissive human tissue culture the MVA deletion mutant viruses produced reduced levels of the VACV envelope antigen B5. Upon mousepox challenge at three weeks after vaccination, MVA-ΔF1L and MVA-ΔE3L exhibited reduced protective capacity in comparison to wildtype MVA. Surprisingly, however, all vaccines proved equally protective against a lethal ECTV infection at two days after vaccination. Accordingly, the deletion mutant MVA vaccines induced high levels of virus-specific CD8+ T cells previously shown to be essential for rapidly protective MVA vaccination. These results suggest that inactivation of the anti-apoptotic genes F1L or E3L modulates the protective capacity of MVA vaccination most likely through the induction of distinct orthopoxvirus specific immunity in the absence of these viral regulatory proteins.

  18. Molecular characterisation of two α-esterase genes involving chlorpyrifos detoxification in the diamondback moth, Plutella xylostella.

    Science.gov (United States)

    Xie, Miao; Ren, Na-Na; You, Yan-Chun; Chen, Wei-Jun; Song, Qi-Sheng; You, Min-Sheng

    2017-06-01

    Carboxylesterases (CarEs) are involved in metabolic detoxification of dietary and environmental xenobiotics in insects. However, owing to the complexity of the protein family, the involvement of CarEs in insecticide metabolism in Plutella xylostella has not been fully elucidated. This study aimed to characterise two CarE genes and assess their potential roles in response to chlorpyrifos in P. xylostella. Synergistic tests showed that triphenyl phosphate decreased the resistance of the third-instar larvae to chlorpyrifos. The treatment of the third-instar larvae with chlorpyrifos at the LC 30 dose led to a significant increase in CarE activity. Two CarE cDNAs (Pxae18 and Pxae28) were subsequently sequenced and characterised. Both genes were expressed predominantly in the larval midgut. Most importantly, two CarE genes showed significantly higher expression in the chlorpyrifos-resistant strain than in the susceptible strain. RNAi knockdown of Pxae18 and Pxae28 significantly increased the mortality to chlorpyrifos from 40% in the control to 73.8 and 63.3% respectively. RNAi knockdown of Pxae18 and Pxae28 significantly inhibited detoxification ability and increased the mortality in P. xylostella. The results indicate that these two CarE genes play important roles in the detoxification of chlorpyrifos in P. xylostella. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  19. E3 protein of bovine coronavirus is a receptor-destroying enzyme with acetylesterase activity

    International Nuclear Information System (INIS)

    Vlasak, R.; Luytjes, W.; Leider, J.; Spaan, W.; Palese, P.

    1988-01-01

    In addition to members of the Orthomyxoviridae and Paramyxoviridae, several coronaviruses have been shown to possess receptor-destroying activities. Purified bovine coronavirus (BCV) preparations have an esterase activity which inactivates O-acetylsialic acid-containing receptors on erythrocytes. Diisopropyl fluorophosphate (DFP) completely inhibits this receptor-destroying activity of BCV, suggesting that the viral enzyme is a serine esterase. Treatment of purified BCV with [ 3 H]DFP and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the proteins revealed that the esterase/receptor-destroying activity of BCV is associated with the E3 protein was specifically phosphorylated. This finding suggests that the esterase/receptor-destroying activity of BCV is associated with the E3 protein. Furthermore, treatment of BCV with DFP dramatically reduced its infectivity in a plaque assay. It is assumed that the esterase activity of BCV is required in an early step of virus replication, possible during virus entry or uncoating

  20. E3 protein of bovine coronavirus is a receptor-destroying enzyme with acetylesterase activity

    Energy Technology Data Exchange (ETDEWEB)

    Vlasak, R.; Luytjes, W.; Leider, J.; Spaan, W.; Palese, P.

    1988-12-01

    In addition to members of the Orthomyxoviridae and Paramyxoviridae, several coronaviruses have been shown to possess receptor-destroying activities. Purified bovine coronavirus (BCV) preparations have an esterase activity which inactivates O-acetylsialic acid-containing receptors on erythrocytes. Diisopropyl fluorophosphate (DFP) completely inhibits this receptor-destroying activity of BCV, suggesting that the viral enzyme is a serine esterase. Treatment of purified BCV with (/sup 3/H)DFP and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the proteins revealed that the esterase/receptor-destroying activity of BCV is associated with the E3 protein was specifically phosphorylated. This finding suggests that the esterase/receptor-destroying activity of BCV is associated with the E3 protein. Furthermore, treatment of BCV with DFP dramatically reduced its infectivity in a plaque assay. It is assumed that the esterase activity of BCV is required in an early step of virus replication, possible during virus entry or uncoating.

  1. E3 ubiquitin ligase gene CMPG1-V from Haynaldia villosa L. contributes to powdery mildew resistance in common wheat (Triticum aestivum L.).

    Science.gov (United States)

    Zhu, Yanfei; Li, Yingbo; Fei, Fei; Wang, Zongkuan; Wang, Wei; Cao, Aizhong; Liu, Yuan; Han, Shuang; Xing, Liping; Wang, Haiyan; Chen, Wei; Tang, Sanyuan; Huang, Xiahe; Shen, Qianhua; Xie, Qi; Wang, Xiue

    2015-10-01

    Powdery mildew is one of the most devastating wheat fungal diseases. A diploid wheat relative, Haynaldia villosa L., is highly resistant to powdery mildew, and its genetic resource of resistances, such as the Pm21 locus, is now widely used in wheat breeding. Here we report the cloning of a resistance gene from H. villosa, designated CMPG1-V, that encodes a U-box E3 ubiquitin ligase. Expression of the CMPG1-V gene was induced in the leaf and stem of H. villosa upon inoculation with Blumeria graminis f. sp. tritici (Bgt) fungus, and the presence of Pm21 is essential for its rapid induction of expression. CMPG1-V has conserved key residues for E3 ligase, and possesses E3 ligase activity in vitro and in vivo. CMPG1-V is localized in the nucleus, endoplasmic reticulum, plasma membrane and partially in trans-Golgi network/early endosome vesicles. Transgenic wheat over-expressing CMPG1-V showed improved broad-spectrum powdery mildew resistance at seedling and adult stages, associated with an increase in expression of salicylic acid-responsive genes, H2 O2 accumulation, and cell-wall protein cross-linking at the Bgt infection sites, and the expression of CMPG1-V in H. villosa was increased when treated with salicylic acid, abscisic acid and H2 O2 . These results indicate the involvement of E3 ligase in defense responses to Bgt fungus in wheat, particularly in broad-spectrum disease resistance, and suggest association of reactive oxidative species and the phytohormone pathway with CMPG1-V-mediated powdery mildew resistance. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

  2. Targeted In Situ Gene Correction of Dysfunctional APOE Alleles to Produce Atheroprotective Plasma ApoE3 Protein

    Directory of Open Access Journals (Sweden)

    Ioannis Papaioannou

    2012-01-01

    Full Text Available Cardiovascular disease is the leading worldwide cause of death. Apolipoprotein E (ApoE is a 34-kDa circulating glycoprotein, secreted by the liver and macrophages with pleiotropic antiatherogenic functions and hence a candidate to treat hypercholesterolaemia and atherosclerosis. Here, we describe atheroprotective properties of ApoE, though also potential proatherogenic actions, and the prevalence of dysfunctional isoforms, outline conventional gene transfer strategies, and then focus on gene correction therapeutics that can repair defective APOE alleles. In particular, we discuss the possibility and potential benefit of applying in combination two technical advances to repair aberrant APOE genes: (i an engineered endonuclease to introduce a double-strand break (DSB in exon 4, which contains the common, but dysfunctional, ε2 and ε4 alleles; (ii an efficient and selectable template for homologous recombination (HR repair, namely, an adeno-associated viral (AAV vector, which harbours wild-type APOE sequence. This technology is applicable ex vivo, for example to target haematopoietic or induced pluripotent stem cells, and also for in vivo hepatic gene targeting. It is to be hoped that such emerging technology will eventually translate to patient therapy to reduce CVD risk.

  3. E3 Staff Database

    Data.gov (United States)

    US Agency for International Development — E3 Staff database is maintained by E3 PDMS (Professional Development & Management Services) office. The database is Mysql. It is manually updated by E3 staff as...

  4. Overexpression of E3 Ubiquitin Ligase Gene AdBiL Contributes to Resistance against Chilling Stress and Leaf Mold Disease in Tomato

    Directory of Open Access Journals (Sweden)

    Shuangchen Chen

    2017-06-01

    Full Text Available Ubiquitination is a common regulatory mechanism, playing a critical role in diverse cellular and developmental processes in eukaryotes. However, a few reports on the functional correlation between E3 ubiquitin ligases and reactive oxygen species (ROS or reactive nitrogen species (RNS metabolism in response to stress are currently available in plants. In the present study, the E3 ubiquitin ligase gene AdBiL (Adi3 Binding E3 Ligase was introduced into tomato line Ailsa Craig via Agrobacterium-mediated method. Transgenic lines were confirmed for integration into the tomato genome using PCR. Transcription of AdBiL in various transgenic lines was determined using real-time PCR. Evaluation of stress tolerance showed that T1 generation of transgenic tomato lines showed only mild symptoms of chilling injury as evident by higher biomass accumulation and chlorophyll content than those of non-transformed plants. Compared with wild-type plants, the contents of AsA, AsA/DHA, GSH and the activity of GaILDH, γ-GCS and GSNOR were increased, while H2O2, O2.−, MDA, NO, SNOs, and GSNO accumulations were significantly decreased in AdBiL overexpressing plants in response to chilling stress. Furthermore, transgenic tomato plants overexpressing AdBiL showed higher activities of enzymes such as G6PDH, 6PGDH, NADP-ICDH, and NADP-ME involved in pentose phosphate pathway (PPP. The transgenic tomato plants also exhibited an enhanced tolerance against the necrotrophic fungus Cladosporium fulvum. Tyrosine nitration protein was activated in the plants infected with leaf mold disease, while the inhibition could be recovered in AdBiL gene overexpressing lines. Taken together, our results revealed a possible physiological role of AdBiL in the activation of the key enzymes of AsA–GSH cycle, PPP and down-regulation of GSNO reductase, thereby reducing oxidative and nitrosative stress in plants. This study demonstrates an optimized transgenic strategy using AdBiL gene for crop

  5. A feruloyl esterase derived from a leachate metagenome library

    CSIR Research Space (South Africa)

    Rashamuse, K

    2012-01-01

    Full Text Available A feruloyl esterase encoding gene (designated fae6), derived from a leachate metagenomic library, was cloned and the nucleotide sequence of the insert DNA determined. Translational analysis revealed that fae6 consists of a 515 amino acid polypeptide...

  6. The constitutive activation of the CEF-4/9E3 chemokine gene depends on C/EBPbeta in v-src transformed chicken embryo fibroblasts

    DEFF Research Database (Denmark)

    Gagliardi, M; Maynard, S; Bojovic, B

    2001-01-01

    The CEF-4/9E3 chemokine gene is expressed constitutively in chicken embryo fibroblasts (CEF) transformed by the Rous sarcoma virus (RSV). This aberrant induction is controlled at the transcriptional and post-transcriptional levels. Transcriptional activation depends on multiple elements of the CEF....../EBPbeta binds to a second element located in proximity of the TRE. A mutation of this distal CAAT box impaired the activation of the CEF-4 promoter by pp60(v-src) indicating that this element is also part of the SRU. Using the RCASBP retroviral vector, we expressed a dominant negative mutant of C....../EBPbeta (designated Delta184-C/EBPbeta) in RSV-transformed CEF. Delta184-C/EBPbeta decreased the accumulation of the CEF-4 mRNA and activation of the CEF-4 promoter by pp60(v-src). The induction of the Cox-2 gene (CEF-147) was also reduced by Delta184-C/EBPbeta. The effect of the dominant negative mutant was observed...

  7. The SUD1 gene encodes a putative E3 ubiquitin ligase and is a positive regulator of 3-hydroxy-3-methylglutaryl coenzyme a reductase activity in Arabidopsis.

    Science.gov (United States)

    Doblas, Verónica G; Amorim-Silva, Vítor; Posé, David; Rosado, Abel; Esteban, Alicia; Arró, Montserrat; Azevedo, Herlander; Bombarely, Aureliano; Borsani, Omar; Valpuesta, Victoriano; Ferrer, Albert; Tavares, Rui M; Botella, Miguel A

    2013-02-01

    The 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) enzyme catalyzes the major rate-limiting step of the mevalonic acid (MVA) pathway from which sterols and other isoprenoids are synthesized. In contrast with our extensive knowledge of the regulation of HMGR in yeast and animals, little is known about this process in plants. To identify regulatory components of the MVA pathway in plants, we performed a genetic screen for second-site suppressor mutations of the Arabidopsis thaliana highly drought-sensitive drought hypersensitive2 (dry2) mutant that shows decreased squalene epoxidase activity. We show that mutations in SUPPRESSOR OF DRY2 DEFECTS1 (SUD1) gene recover most developmental defects in dry2 through changes in HMGR activity. SUD1 encodes a putative E3 ubiquitin ligase that shows sequence and structural similarity to yeast Degradation of α factor (Doα10) and human TEB4, components of the endoplasmic reticulum-associated degradation C (ERAD-C) pathway. While in yeast and animals, the alternative ERAD-L/ERAD-M pathway regulates HMGR activity by controlling protein stability, SUD1 regulates HMGR activity without apparent changes in protein content. These results highlight similarities, as well as important mechanistic differences, among the components involved in HMGR regulation in plants, yeast, and animals.

  8. Xylella fastidiosa esterase rather than hydroxynitrile lyase.

    Science.gov (United States)

    Torrelo, Guzman; Ribeiro de Souza, Fayene Zeferino; Carrilho, Emanuel; Hanefeld, Ulf

    2015-03-02

    In 2009, we reported that the product of the gene SCJ21.16 (XFa0032) from Xylella fastidiosa, a xylem-restricted plant pathogen that causes a range of diseases in several important crops, encodes a protein (XfHNL) with putative hydroxynitrile lyase activity. Sequence analysis and activity tests indicated that XfHNL exhibits an α/β-hydrolase fold and could be classified as a member of the family of FAD-independent HNLs. Here we provide a more detailed sequence analysis and new experimental data. Using pure heterologously expressed XfHNL we show that this enzyme cannot catalyse the cleavage/synthesis of mandelonitrile and that this protein is in fact a non-enantioselective esterase. Homology modelling and ligand docking simulations were used to study the active site and support these results. This finding could help elucidate the common ancestor of esterases and hydroxynitrile lyases with an α/β -hydrolase fold. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Reversal of hypercholesterolemia in apolipoprotein E2 and apolipoprotein E3-Leiden transgenic mice by adenovirus-mediated gene transfer of the VLDL receptor

    NARCIS (Netherlands)

    Dijk, K.W. van; Vlijmen, B.J.M. van; Zee, A. van der; Hof, B. van 't; Boom, H. van der; Kobayashi, K.; Chan, L.; Havekes, L.M.; Hofker, M.H.

    1998-01-01

    We have investigated the interaction of apolipoprotein E2(Arg158- Cys) (apoE2) and apolipoprotein E3Leiden (apoE3-Leiden) with the very low density lipoprotein (VLDL) receptor in vivo and in vitro to define the possible role of this receptor in lipoprotein metabolism and atherosclerosis. The in vivo

  10. Analyses of Evolutionary Characteristics of the Hemagglutinin-Esterase Gene of Influenza C Virus during a Period of 68 Years Reveals Evolutionary Patterns Different from Influenza A and B Viruses

    Directory of Open Access Journals (Sweden)

    Yuki Furuse

    2016-11-01

    Full Text Available Infections with the influenza C virus causing respiratory symptoms are common, particularly among children. Since isolation and detection of the virus are rarely performed, compared with influenza A and B viruses, the small number of available sequences of the virus makes it difficult to analyze its evolutionary dynamics. Recently, we reported the full genome sequence of 102 strains of the virus. Here, we exploited the data to elucidate the evolutionary characteristics and phylodynamics of the virus compared with influenza A and B viruses. Along with our data, we obtained public sequence data of the hemagglutinin-esterase gene of the virus; the dataset consists of 218 unique sequences of the virus collected from 14 countries between 1947 and 2014. Informatics analyses revealed that (1 multiple lineages have been circulating globally; (2 there have been weak and infrequent selective bottlenecks; (3 the evolutionary rate is low because of weak positive selection and a low capability to induce mutations; and (4 there is no significant positive selection although a few mutations affecting its antigenicity have been induced. The unique evolutionary dynamics of the influenza C virus must be shaped by multiple factors, including virological, immunological, and epidemiological characteristics.

  11. Third-Generation Sequencing and Analysis of Four Complete Pig Liver Esterase Gene Sequences in Clones Identified by Screening BAC Library.

    Science.gov (United States)

    Zhou, Qiongqiong; Sun, Wenjuan; Liu, Xiyan; Wang, Xiliang; Xiao, Yuncai; Bi, Dingren; Yin, Jingdong; Shi, Deshi

    2016-01-01

    Pig liver carboxylesterase (PLE) gene sequences in GenBank are incomplete, which has led to difficulties in studying the genetic structure and regulation mechanisms of gene expression of PLE family genes. The aim of this study was to obtain and analysis of complete gene sequences of PLE family by screening from a Rongchang pig BAC library and third-generation PacBio gene sequencing. After a number of existing incomplete PLE isoform gene sequences were analysed, primers were designed based on conserved regions in PLE exons, and the whole pig genome used as a template for Polymerase chain reaction (PCR) amplification. Specific primers were then selected based on the PCR amplification results. A three-step PCR screening method was used to identify PLE-positive clones by screening a Rongchang pig BAC library and PacBio third-generation sequencing was performed. BLAST comparisons and other bioinformatics methods were applied for sequence analysis. Five PLE-positive BAC clones, designated BAC-10, BAC-70, BAC-75, BAC-119 and BAC-206, were identified. Sequence analysis yielded the complete sequences of four PLE genes, PLE1, PLE-B9, PLE-C4, and PLE-G2. Complete PLE gene sequences were defined as those containing regulatory sequences, exons, and introns. It was found that, not only did the PLE exon sequences of the four genes show a high degree of homology, but also that the intron sequences were highly similar. Additionally, the regulatory region of the genes contained two 720bps reverse complement sequences that may have an important function in the regulation of PLE gene expression. This is the first report to confirm the complete sequences of four PLE genes. In addition, the study demonstrates that each PLE isoform is encoded by a single gene and that the various genes exhibit a high degree of sequence homology, suggesting that the PLE family evolved from a single ancestral gene. Obtaining the complete sequences of these PLE genes provides the necessary foundation for

  12. Esterase profile of human masseter muscle

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D; Vilmann, H

    1988-01-01

    The esterase profile of fresh human masseter muscle was investigated by use of histochemistry and electrophoresis. The histochemical methods included reactions for alpha-naphthyl esterase, myofibrillar ATPase, reverse myofibrillar ATPase and succinic dehydrogenase. In frozen sections of the muscle...... the coloured reaction product for esterases was present both as a diffuse sarcoplasmic coloration and as distinct granules. The intensity of diffuse reaction was used to classify the muscle fibres as strongly, moderately and weakly reacting. The fibres with strong esterase activity belonged to Type I and ii......C. iM and Type II A fibres showed a moderate esterase reaction and Type II B fibres had a low activity. The electrophoretic gels stained for esterase activity showed that the human masseter muscle possesses a slow migrating double band with high enzyme activity and a cascade of faster migrating...

  13. Effect of halogenated benzenes on acetanilide esterase, acetanilide hydroxylase and procaine esterase in rats.

    Science.gov (United States)

    Carlson, G P; Dziezak, J D; Johnson, K M

    1979-07-01

    1,2,4-Trichlorobenzene, 1,3,5-trichlorobenzene, hexachlorobenzene, 1,2,4-tribromobenzene, 1,3,5-tribromobenzene and hexabromobenzene were compared for their abilities to induce acetanilide esterase, acentailide hydroxylase and procaine esterase. Except for hexabromobenzene all induced acetanilide esterase whereas the hydroxylation of acetanilide was seen only with the fully halogenated benzenes and with 1,3,5-tribromobenzene. Hepatic procaine esterase activity was increased by the three chlorinated benzenes and 1,2,4-tribromobenzene.

  14. Overexpression of esterase D in kidney from trisomy 13 fetuses

    Energy Technology Data Exchange (ETDEWEB)

    Loughna, S.; Moore, G. (Institute of Obstetrics and Gynaecology, London (United Kingdom)); Gau, G.; Blunt, S. (Cytogenetics Lab., London (United Kingdom)); Nicolaides, K. (King' s College School of Medicine and Dentistry, London (United Kingdom))

    1993-10-01

    Human trisomy 13 (Patau syndrome) occurs in approximately 1 in 5,000 live births. It is compatible with life, but prolonged survival is rare. Anomalies often involve the urogenital, cardiac, craniofacial, and central nervous systems. It is possible that these abnormalities may be due to the overexpression of developmentally important genes on chromosome 13. The expression of esterase D (localized to chromosome 13q14.11) has been investigated in both muscle and kidney from trisomy 13 fetuses and has been compared with normal age- and sex-matched fetal tissues, by using northern analysis. More than a twofold increase in expression of esterase D was found in the kidney of two trisomy 13 fetuses, with normal levels in a third. Overexpression was not seen in the muscle tissues from these fetuses. 34 refs., 3 figs., 2 tabs.

  15. Esterase reactions in acute myelomonocytic leukemia.

    Science.gov (United States)

    Kass, L

    1977-05-01

    Specific and nonspecific esterase reactions of bone marrow cells from 14 patients with untreated acute myelomonocytic leukemia and six patients with acute histiomonocytic leukemia were examined. The technic for esterase determination permitted simultaneous visualization of both esterases on the same glass coverslip containing the marrow cells. In cases of acute histiomonocytic leukemia, monocytes, monocytoid hemohistioblasts and undifferentiated blasts stained intensely positive for nonspecific esterase, using alpha-naphthyl acetate as the substrate. No evidence of specific esterase activity using naphthol ASD-chloroacetate as the substrate and fast blue BBN as the dye coupler was apparent in these cells. In all of the cases of acute myelomonocytic leukemia, both specific and nonspecific esterases were visualized within monocytes, monocytoid cells, and granulocytic cells that had monocytoid-type nuclei. Nonspecific esterase activity was not observed in polymorphonuclear leukocytes in cases of myelomonocytic leukemia. The results support a current viewpoint that acute myelomonocytic leukemia may be a variant of acute myeloblastic leukemia, and that cytochemically, many of the leukemic cells in myelomonocytic leukemia share properties of both granulocytes and monocytes.

  16. Esterase resistant to inactivation by heavy metals

    KAUST Repository

    El, Dorry Hamza

    2014-09-25

    EstATII is an esterase that a halotolerant, thermophilic and resistant to a spectrum of heavy metals including toxic concentration of metals. It was isolated from the lowest convective layer of the Atlantis II Red Sea brine pool. The Atlantis II brine pool is an extreme environment that possesses multiple harsh conditions such as; high temperature, salinity, pH and high concentration of metals, including toxic heavy metals. A fosmid metagenomic library using DNA isolated from the lowest convective layer this pool was used to identify EstATII. Polynucleotides encoding EstATII and similar esterases are disclosed and can be used to make EstATII. EstATII or compositions or apparatuses that contain it may be used in various processes employing lipases/esterases especially when these processes are performed under harsh conditions that inactivate other kinds of lipases or esterases.

  17. Effects of Model Salivary Esterases and MMP Inhibition on the Restoration's Marginal Integrity and Potential Degradative Contribution of Cariogenic Bacteria

    Science.gov (United States)

    Huang, Bo

    Enzyme-catalyzed degradation of the restoration-tooth interface compromises interfacial integrity, thereby contributing to secondary caries, which is a major cause of resin-based restoration failure. It is hypothesized that in addition to salivary esterases, the cariogenic bacterium Streptococcus mutans has specific esterases that degrade the resin-dentin interface, releasing biodegradation by- products (BBPs) such as bis-hydroxy-propoxy-phenyl-propane (BisHPPP). In turn, BisHPPP affects S. mutans by stimulating the expression of esterases. Another hypothesis is that the biostability of the resin-dentin interface is affected by simulated salivary esterases, dentinal matrix metalloproteinase (MMP) inhibition, and restorative materials. To test the first hypothesis, putative esterase genes in S. mutans UA159 were identified, purified, and characterized. SMU_118c was identified as the dominant esterase in S. mutans UA159 and showed a similar hydrolytic activity profile to salivary esterases. BisHPPP upregulated expression of the SMU_118c gene and related protein in a concentration-dependent manner. This positive feedback process could accelerate the degradation of the restoration-tooth interface and lead to premature restoration failure. To test the second hypothesis, an in vitro model was established to evaluate the effects of salivary esterases, MMP inhibition and restorative materials on interfacial integrity. It was confirmed that interfacial integrity was compromised with time and was further deteriorated by simulated salivary esterases, as indicated by the greater depth of bacterial ingress and more bacterial biomass of biofilm along the interface. However, this process could be modulated by using different restorative materials and MMPs inhibition. This project elucidated the mechanistic interaction between oral bacteria and restorative materials and established a new, in vitro, and physiologically relevant model to assess the effect of material chemistry

  18. Isolation and characterization of novel lipases/esterases from a bovine rumen metagenome.

    Science.gov (United States)

    Privé, Florence; Newbold, C Jamie; Kaderbhai, Naheed N; Girdwood, Susan G; Golyshina, Olga V; Golyshin, Peter N; Scollan, Nigel D; Huws, Sharon A

    2015-07-01

    Improving the health beneficial fatty acid content of meat and milk is a major challenge requiring an increased understanding of rumen lipid metabolism. In this study, we isolated and characterized rumen bacterial lipases/esterases using functional metagenomics. Metagenomic libraries were constructed from DNA extracted from strained rumen fluid (SRF), solid-attached bacteria (SAB) and liquid-associated rumen bacteria (LAB), ligated into a fosmid vector and subsequently transformed into an Escherichia coli host. Fosmid libraries consisted of 7,744; 8,448; and 7,680 clones with an average insert size of 30 to 35 kbp for SRF, SAB and LAB, respectively. Transformants were screened on spirit blue agar plates containing tributyrin for lipase/esterase activity. Five SAB and four LAB clones exhibited lipolytic activity, and no positive clones were found in the SRF library. Fosmids from positive clones were pyrosequenced and twelve putative lipase/esterase genes and two phospholipase genes retrieved. Although the derived proteins clustered into diverse esterase and lipase families, a degree of novelty was seen, with homology ranging from 40 to 78% following BlastP searches. Isolated lipases/esterases exhibited activity against mostly short- to medium-chain substrates across a range of temperatures and pH. The function of these novel enzymes recovered in ruminal metabolism needs further investigation, alongside their potential industrial uses.

  19. Three feruloyl esterases in Cellulosilyticum ruminicola H1 act synergistically to hydrolyze esterified polysaccharides.

    Science.gov (United States)

    Li, Jiabao; Cai, Shichun; Luo, Yuanming; Dong, Xiuzhu

    2011-09-01

    Feruloyl esterases (Faes) constitute a subclass of carboxyl esterases that specifically hydrolyze the ester linkages between ferulate and polysaccharides in plant cell walls. Until now, the described microbial Faes were mainly from fungi. In this study, we report that Cellulosilyticum ruminicola H1, a previously described fibrolytic rumen bacterium, possesses three different active feruloyl esterases, FaeI, FaeII, and FaeIII. Phylogenetic analysis classified the described bacterial Faes into two types, FaeI and FaeII in type I and FaeIII in type II. Substrate specificity assays indicated that FaeI is more active against the ester bonds in natural hemicelluloses and FaeIII preferentially attacks the ferulate esters with a small moiety, such as methyl groups, while FaeII is active on both types of substrates. Among the three feruloyl esterase genes, faeI was the only one induced significantly by xylose and xylan, while pectin appeared to moderately induce the three genes during the late log phase to stationary phase. Western blot analysis determined that FaeI and FaeIII were secreted and cytoplasmic proteins, respectively, whereas FaeII seemed to be cell associated. The addition of FaeI and FaeII but not FaeIII enhanced the activity of a xylanase on maize cob, suggesting a synergy of the former two with xylanase. Hence, we propose that the three feruloyl esterases work in concert to hydrolyze ferulate esters in natural hemicelluloses.

  20. Genome-wide RNAi screen reveals the E3 SUMO-protein ligase gene SIZ1 as a novel determinant of furfural tolerance in Saccharomyces cerevisiae

    OpenAIRE

    Xiao, Han; Zhao, Huimin

    2014-01-01

    Background Furfural is a major growth inhibitor in lignocellulosic hydrolysates and improving furfural tolerance of microorganisms is critical for rapid and efficient fermentation of lignocellulosic biomass. In this study, we used the RNAi-Assisted Genome Evolution (RAGE) method to select for furfural resistant mutants of Saccharomyces cerevisiae, and identified a new determinant of furfural tolerance. Results By using a genome-wide RNAi (RNA-interference) screen in S. cerevisiae for genes in...

  1. Esterase variation in Turkish white-toothed shrews (Crocidura: Record of a trimeric esterase

    Directory of Open Access Journals (Sweden)

    Tez C.

    2009-01-01

    Full Text Available This study focuses on esterase variation of the genus Crocidura in Turkey. A total of 248 white-toothed shrews were analyzed by means of cellulose acetate gel electrophoresis. Liver tissue and alfa naphthyl acetate were used to investigate esterase variation in Turkish white-toothed shrews. A different esterase banding pattern was found in one Crocidura individual. This phenotype had four anodally migrated bands on cellulose acetate gel. The Crocidura individual displaying the given phenotype was identified as Crocidura suaveolens. The different esterase banding pattern observed in this study is considered to be a result of the trimeric structure of esterase in the lesser white-toothed shrew (Crocidura suaveolens.

  2. Bag1 Co-chaperone Promotes TRC8 E3 Ligase-dependent Degradation of Misfolded Human Ether a Go-Go-related Gene (hERG) Potassium Channels.

    Science.gov (United States)

    Hantouche, Christine; Williamson, Brittany; Valinsky, William C; Solomon, Joshua; Shrier, Alvin; Young, Jason C

    2017-02-10

    Cardiac long QT syndrome type 2 is caused by mutations in the human ether a go-go-related gene (hERG) potassium channel, many of which cause misfolding and degradation at the endoplasmic reticulum instead of normal trafficking to the cell surface. The Hsc70/Hsp70 chaperones assist the folding of the hERG cytosolic domains. Here, we demonstrate that the Hsp70 nucleotide exchange factor Bag1 promotes hERG degradation by the ubiquitin-proteasome system at the endoplasmic reticulum to regulate hERG levels and channel activity. Dissociation of hERG complexes containing Hsp70 and the E3 ubiquitin ligase CHIP requires the interaction of Bag1 with Hsp70, but this does not involve the Bag1 ubiquitin-like domain. The interaction with Bag1 then shifts hERG degradation to the membrane-anchored E3 ligase TRC8 and its E2-conjugating enzyme Ube2g2, as determined by siRNA screening. TRC8 interacts through the transmembrane region with hERG and decreases hERG functional expression. TRC8 also mediates degradation of the misfolded hERG-G601S disease mutant, but pharmacological stabilization of the mutant structure prevents degradation. Our results identify TRC8 as a previously unknown Hsp70-independent quality control E3 ligase for hERG. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Schmidt Reaction of E-3-Benzylidenechromanones and E-3-Benzylidenethiochromanones

    Directory of Open Access Journals (Sweden)

    Tapas K. Mandal

    2013-01-01

    Full Text Available On treatment with NaN3/c. H2SO4-HOAc or NaN3/TFA, E-3-benzylidenechromanones are mostly converted to E-β-aminobenzylidenechromanones while E-3-benzylidenethiochromanones are converted to 3-benzoylthiochromones. A structurally new type of product has been isolated for the reaction of E-3-benzylidene-4′-methoxychromanone with NaN3/TFA. Mechanistic paths have been suggested for formation of the products.

  4. Metagenomic mining of feruloyl esterases from termite enteric flora

    CSIR Research Space (South Africa)

    Rashamuse, K

    2014-01-01

    Full Text Available A metagenome expression library was created from Trinervitermes trinervoides termite hindgut symbionts and subsequently screened for feruloyl esterase (FAE) activities, resulting in seven recombinant fosmids conferring feruloyl esterase phenotypes...

  5. Esterases of laboratory-reared and field-collected cotton boll weevils, Anthonomus grandis Boh.: polymorphism of adult esterases and formal genetics of esterase II.

    Science.gov (United States)

    Biggers, C J; Bancroft, H R

    1977-04-01

    The esterases of the cotton boll weevil were separated by polyacrylamide gel electrophoresis into four major regions. These were named Est I-IV in order of migration from anode to origin. Polymorphism was observed in all regions. The Est II region was shown to consist of no more than two bands (fast and slow). The inheritance of the fast and slow bands of Est II was demonstrated to be controlled by codominant autosomal alleles. Analysis of the gene frequency of the Est II region showed that one field population was consistent with the Hardy-Weinberg law (P = 0.995), while a second field population was not at equilibrium (P less than 0.001).

  6. The effect of fixation on esterases

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D

    1984-01-01

    The localization of reaction product for non-specific esterase from fresh and aldehyde treated glandular tissue was examined. The electrophoretical studies showed a selective inhibition of certain isoenzymes and a change in mobility of some bands caused by aldehyde fixation. In sections a granular...

  7. Esterase resistant to inactivation by heavy metals

    KAUST Repository

    El, Dorry Hamza; Siam, Rania; Mohamed, Yasmine M.

    2014-01-01

    EstATII is an esterase that a halotolerant, thermophilic and resistant to a spectrum of heavy metals including toxic concentration of metals. It was isolated from the lowest convective layer of the Atlantis II Red Sea brine pool. The Atlantis II

  8. Cholesterol esterase activity of human intestinal mucosa

    International Nuclear Information System (INIS)

    Ponz de Leon, M.; Carubbi, F.; Di Donato, P.; Carulli, N.

    1985-01-01

    It has been suggested that cholesterol absorption in humans is dependent on bile acid pool composition and that expansion of the cholic acid pool size is followed by an increase of the absorption values. Similar observations were reported in rats. In the present study, therefore, the authors investigated some general properties of human intestinal cholesterol esterase, with particular emphasis on the effect of bile acids on this enzymatic activity. Twenty-nine segments of small intestine were taken during operations; the enzymatic activity was studied by using mucosal homogenate as a source of enzyme and oleic acid, cholesterol, and 14 C-labeled cholesterol as substrates. The time-activity relationship was linear within the first two hours; optimal pH for esterification ranged between 5 and 6.2. There was little difference between the esterifying activity of the jejunal and ileal mucosa. Esterification of cholesterol was observed with all the investigated fatty acids but was maximal with oleic acid. Bile acids did not affect cholesterol esterase activity when present in the incubation mixture at 0.1 and 1.0 mM; the enzymatic activity, however, was significantly inhibited when bile acids were added at 20 mM. In conclusion, this study has shown that the human intestinal mucosa possesses a cholesterol esterase activity; at variance with the rat, however, the human enzyme does not seem to be stimulated by trihydroxy bile acids

  9. Non-specific esterases in partly mineralized bovine enamel

    DEFF Research Database (Denmark)

    Moe, D; Kirkeby, S

    1990-01-01

    Activity for non-specific esterase was demonstrated in the matrix of developing bovine enamel with alpha-naphthyl acetate and 5-bromoindoxyl acetate as the esterase substrates. By use of high-performance liquid chromatography gel filtration, ion-exchange chromatography, and electrophoresis three...... esterases were shown to be present in the enamel matrix. The enzymes showed highest activity at pH 6.5-7.5. In sections a strong reaction was observed in the secretory ameloblasts. The esterases may be proteolytic enzymes that participate in the degradation of the matrix proteins....

  10. A p-coumaroyl esterase from Rhizoctonia solani with a pronounced chlorogenic acid esterase activity.

    Science.gov (United States)

    Nieter, Annabel; Kelle, Sebastian; Linke, Diana; Berger, Ralf G

    2017-07-25

    Extracellular esterase activity was detected in submerged cultures of Rhizoctonia solani grown in the presence of sugar beet pectin or Tween 80. Putative type B feruloyl esterase (FAE) coding sequences found in the genome data of the basidiomycete were heterologously expressed in Pichia pastoris. Recombinant enzyme production on the 5-L bioreactor scale (Rs pCAE: 3245UL -1 ) exceeded the productivity of the wild type strain by a factor of 800. Based on substrate specificity profiling, the purified recombinant Rs pCAE was classified as a p-coumaroyl esterase (pCAE) with a pronounced chlorogenic acid esterase side activity. The Rs pCAE was also active on methyl cinnamate, caffeate and ferulate and on feruloylated saccharides. The unprecedented substrate profile of Rs pCAE together with the lack of sequence similarity to known FAEs or pCAEs suggested that the Rs pCAE represents a new type of enzyme. Hydroxycinnamic acids were released from agro-industrial side-streams, such as destarched wheat bran (DSWB), sugar beet pectin (SBP) and coffee pulp (CP). Overnight incubation of coffee pulp with the Rs pCAE resulted in the efficient release of p-coumaric (100%), caffeic (100%) and ferulic acid (85%) indicating possible applications for the valorization of food processing wastes and for the enhanced degradation of lignified biomass. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Cloning, expression and characterization of a novel esterase from a South China Sea sediment metagenome

    Science.gov (United States)

    Zhang, Hao; Li, Fuchao; Chen, Huaxin; Zhao, Jin; Yan, Jinfei; Jiang, Peng; Li, Ronggui; Zhu, Baoli

    2015-07-01

    Lipolytic enzymes, including esterases and lipases, represent a group of hydrolases that catalyze the cleavage and formation of ester bonds. A novel esterase gene, scsEst01, was cloned from a South China Sea sediment metagenome. The scsEst01 gene consisted of 921 bp encoding 307 amino acid residues. The predicted amino acid sequence shared less than 90% identity with other lipolytic enzymes in the NCBI nonredundant protein database. ScsEst01 was successfully co-expressed in Escherichia coli BL21 (DE3) with chaperones (dnaK-dnaJ-grpE) to prevent the formation of inclusion bodies. The recombinant protein was purified on an immobilized metal ion affinity column containing chelating Sepharose charged with Ni2+. The enzyme was characterized using p -nitrophenol butyrate as a substrate. ScsEst01 had the highest lipolytic activity at 35°C and pH 8.0, indicative of a meso-thermophilic alkaline esterase. ScsEst01 was thermostable at 20°C. The lipolytic activity of scsEst01 was strongly increased by Fe2+, Mn2+ and 1% Tween 80 or Tween 20.

  12. A Novel Halotolerant Thermoalkaliphilic Esterase from Marine Bacterium Erythrobacter seohaensis SW-135

    Directory of Open Access Journals (Sweden)

    Ying-Yi Huo

    2017-11-01

    Full Text Available A novel esterase gene, e69, was cloned from Erythrobacter seohaensis SW-135, which was isolated from a tidal flat sediment of the Yellow Sea in Korea. This gene is 825 bp in length and codes for a 29.54 kDa protein containing 274 amino acids. Phylogenetic analysis showed that E69 is a new member of the bacterial lipolytic enzyme family IV. This enzyme exhibited the highest level of activity toward p-nitrophenyl (NP butyrate but little or no activity toward the other p-NP esters tested. The optimum temperature and pH of the catalytic activity of E69 were 60°C and pH 10.5, respectively. The enzyme exhibited stable activity over a wide range of alkaline pH values (7.5–9.5. In addition, E69 was found to be a halotolerant esterase as it exhibited the highest hydrolytic activity in the presence of 0.5 M NaCl and was still active in the presence of 3 M NaCl. Moreover, it possessed some degree of tolerance to Triton X-100 and several organic solvents. Through homology modeling and comparison with other esterases, it was suggested that the absence of the cap domain and its narrow substrate-binding pocket might be responsible for its narrow substrate specificity. Sequence and structural analysis results suggested that its high ratio of negatively to positively charged residues, large hydrophobic surface area, and negative electrostatic potential on the surface may be responsible for its alkaline adaptation. The results of this study provide insight into marine alkaliphilic esterases, and the unique properties of E69 make it a promising candidate as a biocatalyst for industrial applications.

  13. Novel Cold-Adapted Esterase MHlip from an Antarctic Soil Metagenome

    Directory of Open Access Journals (Sweden)

    Moreno Galleni

    2013-01-01

    Full Text Available An Antarctic soil metagenomic library was screened for lipolytic enzymes and allowed for the isolation of a new cytosolic esterase from the a/b hydrolase family 6, named MHlip. This enzyme is related to hypothetical genes coding esterases, aryl-esterases and peroxydases, among others. MHlip was produced, purified and its activity was determined. The substrate profile of MHlip reveals a high specificity for short p-nitrophenyl-esters. The apparent optimal activity of MHlip was measured for p-nitrophenyl-acetate, at 33 °C, in the pH range of 6–9. The MHlip thermal unfolding was investigated by spectrophotometric methods, highlighting a transition (Tm at 50 °C. The biochemical characterization of this enzyme showed its adaptation to cold temperatures, even when it did not present evident signatures associated with cold-adapted proteins. Thus, MHlip adaptation to cold probably results from many discrete structural modifications, allowing the protein to remain active at low temperatures. Functional metagenomics is a powerful approach to isolate new enzymes with tailored biophysical properties (e.g., cold adaptation. In addition, beside the ever growing amount of sequenced DNA, the functional characterization of new catalysts derived from environment is still required, especially for poorly characterized protein families like α/b hydrolases.

  14. Biochemical Characterization of a Family 15 Carbohydrate Esterase from a Bacterial Marine Arctic Metagenome.

    Directory of Open Access Journals (Sweden)

    Concetta De Santi

    Full Text Available The glucuronoyl esterase enzymes of wood-degrading fungi (Carbohydrate Esterase family 15; CE15 form part of the hemicellulolytic and cellulolytic enzyme systems that break down plant biomass, and have possible applications in biotechnology. Homologous enzymes are predicted in the genomes of several bacteria, however these have been much less studied than their fungal counterparts. Here we describe the recombinant production and biochemical characterization of a bacterial CE15 enzyme denoted MZ0003, which was identified by in silico screening of a prokaryotic metagenome library derived from marine Arctic sediment. MZ0003 has high similarity to several uncharacterized gene products of polysaccharide-degrading bacterial species, and phylogenetic analysis indicates a deep evolutionary split between these CE15s and fungal homologs.MZ0003 appears to differ from previously-studied CE15s in some aspects. Some glucuronoyl esterase activity could be measured by qualitative thin-layer chromatography which confirms its assignment as a CE15, however MZ0003 can also hydrolyze a range of other esters, including p-nitrophenyl acetate, which is not acted upon by some fungal homologs. The structure of MZ0003 also appears to differ as it is predicted to have several large loop regions that are absent in previously studied CE15s, and a combination of homology-based modelling and site-directed mutagenesis indicate its catalytic residues deviate from the conserved Ser-His-Glu triad of many fungal CE15s. Taken together, these results indicate that potentially unexplored diversity exists among bacterial CE15s, and this may be accessed by investigation of the microbial metagenome. The combination of low activity on typical glucuronoyl esterase substrates, and the lack of glucuronic acid esters in the marine environment suggest that the physiological substrate of MZ0003 and its homologs is likely to be different from that of related fungal enzymes.

  15. Acidic-alkaline ferulic acid esterase from Chaetomium thermophilum var. dissitum: Molecular cloning and characterization of recombinant enzyme expressed in Pichia pastoris

    DEFF Research Database (Denmark)

    Dotsenko, Gleb; Tong, Xiaoxue; Pilgaard, Bo

    2016-01-01

    A novel ferulic acid esterase encoding gene CtFae, was successfully cloned from a highly esterase active strain of the thermophile ascomycetous fungus Chaetomium thermophilum var. dissitum; the gene was heterologously expressed in Pichia pastoris KM71H. The recombinant enzyme (CtFae) was purified...... to homogeneity and subsequently characterized. CtFae was active towards synthetic esters of ferulic, p-coumaric, and caffeic acids, as well as towards wide range of p-nitrophenyl substrates. Its temperature and pH optima were 55 °C and pH 6.0, respectively. Enzyme rare features were broad pH optimum, high...

  16. Characterization and distribution of esterase activity in activated sludge

    NARCIS (Netherlands)

    Boczar, BA; Forney, LJ; Begley, WM; Larson, RJ; Federle, TW

    2001-01-01

    The location and activity of esterase enzymes in activated Sludge from three Municipal wastewater treatment plants were characterized using model Substrate, and denaturing and nondenaturing polyacrylamide gel electrophoresis (PAGE) Of particulate, freeze thaw (primarily periplasmic enzymes and those

  17. Pectin methyl esterase activity in apple and orange pulps

    International Nuclear Information System (INIS)

    Abdullaev, A.; Djumaev, B.B.; Djumaev, N.B.; Mukhidinov, Z.K.

    2008-01-01

    The results of pectin methyl esterase activity from apple, orange pulp and orange peel depending of ph and temperature are discussed. It's shown that the methyl esterase activity form apple and orange pulps higher in range of temperatures from +37...+60 d ig C . The analysis of dependence of its activity from ph has shown that in both case the enzyme activity increase with increase of ph

  18. Crystallization and preliminary X-ray diffraction analysis of ybfF, a new esterase from Escherichia coli K12

    Energy Technology Data Exchange (ETDEWEB)

    Park, Suk-Youl; Lee, Sang-Hak; Lee, Jieun; Jung, Che-Hun; Kim, Jeong-Sun, E-mail: jsunkim@chonnam.ac.kr [Department of Chemistry and Institute of Basic Sciences, Chonnam National University, Gwangju 500-757 (Korea, Republic of)

    2007-12-01

    The crystallization of ybfF, a new esterase from E. coli, and the collection of diffraction data to 1.1 Å resolution are reported. The product of the recently discovered ybfF gene, which belongs to the esterase family, does not show high sequence similarity to other esterases. To provide the molecular background to the enzymatic mechanism of the ybfF esterase, the ybfF protein from Escherichia coli K12 (Ec-ybfF) was cloned, expressed and purified. The Ec-ybfF protein was crystallized from 60% Tacsimate and 0.1 M bis-Tris propane buffer pH 7.0. Diffraction data were collected to 1.10 Å resolution using synchrotron radiation. The crystal belongs to the orthorhombic space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 66.09, b = 90.71, c = 92.88 Å. With two Ec-ybfF molecules in the asymmetric unit, the crystal volume per unit protein weight is 2.17 Å{sup 3} Da{sup −1}, corresponding to a solvent content of 42%.

  19. Esterase activity as a novel parameter of spore germination in Bacillus anthracis

    International Nuclear Information System (INIS)

    Ferencko, Linda; Cote, Mindy A.; Rotman, Boris

    2004-01-01

    Spores of Bacillus anthracis were shown to produce esterase activity about 4 min after exposure to conventional germinants such as combinations of amino acids and purine ribosides. Neither amino acids nor ribosides alone induce germination and esterase activity. Expression of esterase activity was chloramphenicol resistant, and correlated with loss of spore refractivity, a traditional parameter of early germination. Based on these observations, we hypothesized that esterase activity could be used as a novel parameter for quantifying early events during spore germination. To test this hypothesis, we measured expression of esterase activity under a variety of germinating conditions. Using diacetyl fluorescein as fluorogenic substrate of esterases, we demonstrated that esterase activity was invariably induced whenever spores were triggered by known germinants. Moreover, D-alanine, an inhibitor of L-alanine-mediated germination, was found to significantly inhibit expression of esterase activity. In terms of molecular mechanisms, esterase expression could represent activation of proteases at the onset of spore germination

  20. Biochemical and Structural Analyses of Two Cryptic Esterases in Bacteroides intestinalis and their Synergistic Activities with Cognate Xylanases.

    Science.gov (United States)

    Wefers, Daniel; Cavalcante, Janaina J V; Schendel, Rachel R; Deveryshetty, Jaigeeth; Wang, Kui; Wawrzak, Zdzislaw; Mackie, Roderick I; Koropatkin, Nicole M; Cann, Isaac

    2017-08-04

    Arabinoxylans are constituents of the human diet. Although not utilizable by the human host, they can be fermented by colonic bacteria. The arabinoxylan backbone is decorated with arabinose side chains that may be substituted with ferulic acid, thus limiting depolymerization to fermentable sugars. We investigated the polypeptides encoded by two genes upregulated during growth of the colonic bacterium Bacteroides intestinalis on wheat arabinoxylan. The recombinant proteins, designated BiFae1A and BiFae1B, were functionally assigned esterase activities. Both enzymes were active on acetylated substrates, although each showed a higher ferulic acid esterase activity on methyl-ferulate. BiFae1A showed a catalytic efficiency of 12mM s -1 on para-nitrophenyl-acetate, and on methyl-ferulate, the value was 27 times higher. BiFae1B showed low catalytic efficiencies for both substrates. Furthermore, the two enzymes released ferulic acid from various structural elements, and NMR spectroscopy indicated complete de-esterification of arabinoxylan oligosaccharides from wheat bran. BiFae1A is a tetramer based on the crystal structure, whereas BiFae1B is a dimer in solution based on size exclusion chromatography. The structure of BiFae1A was solved to 1.98Å resolution, and two tetramers were observed in the asymmetric unit. A flexible loop that may act as a hinge over the active site and likely coordinates critical interactions with the substrate was prominent in BiFae1A. Sequence alignments of the esterase domains in BiFae1B with the feruloyl esterase from Clostridium thermocellum suggest that both domains lack the flexible hinge in BiFae1A, an observation that may partly provide a molecular basis for the differences in activities in the two esterases. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Potato tuber pectin structure is influenced by pectin methyl esterase activity and impacts on cooked potato texture

    OpenAIRE

    Ross, Heather A.; Wright, Kathryn M.; McDougall, Gordon J.; Roberts, Alison G.; Chapman, Sean N.; Morris, Wayne L.; Hancock, Robert D.; Stewart, Derek; Tucker, Gregory A.; James, Euan K.; Taylor, Mark A.

    2010-01-01

    Although cooked potato tuber texture is an important trait that influences consumer preference, a detailed understanding of tuber textural properties at the molecular level is lacking. Previous work has identified tuber pectin methyl esterase activity (PME) as a potential factor impacting on textural properties. In this study, tuber PME isoform and gene expression profiles have been determined in potato germplasm with differing textural properties as assessed using an amended wedge fracture m...

  2. Molecular cloning and characterization of a novel pyrethroid-hydrolyzing esterase originating from the Metagenome

    Directory of Open Access Journals (Sweden)

    Liu Yu

    2008-12-01

    Full Text Available Abstract Background Pyrethroids and pyrethrins are widely used insecticides. Extensive applications not only result in pest resistance to these insecticides, but also may lead to environmental issues and human exposure. Numerous studies have shown that very high exposure to pyrethroids might cause potential problems to man and aquatic organisms. Therefore, it is important to develop a rapid and efficient disposal process to eliminate or minimize contamination of surface water, groundwater and agricultural products by pyrethroid insecticides. Bioremediation is considered to be a reliable and cost-effective technique for pesticides abatement and a major factor determining the fate of pyrethroid pesticides in the environment, and suitable esterase is expected to be useful for potential application for detoxification of pyrethroid residues. Soil is a complex environment considered as one of the main reservoirs of microbial diversity on the planet. However, most of the microorganisms in nature are inaccessible as they are uncultivable in the laboratory. Metagenomic approaches provide a powerful tool for accessing novel valuable genetic resources (novel enzymes and developing various biotechnological applications. Results The pyrethroid pesticides residues on foods and the environmental contamination are a public safety concern. Pretreatment with pyrethroid-hydrolyzing esterase has the potential to alleviate the conditions. To this end, a pyrethroid-hydrolyzing esterase gene was successfully cloned using metagenomic DNA combined with activity-based functional screening from soil, sequence analysis of the DNA responsible for the pye3 gene revealed an open reading frame of 819 bp encoding for a protein of 272 amino acid residues. Extensive multiple sequence alignments of the deduced amino acid of Pye3 with the most homologous carboxylesterases revealed moderate identity (45–49%. The recombinant Pye3 was heterologously expressed in E. coli BL21(DE3

  3. Multiple resistance to pirimiphos-methyl and bifenthrin in Tribolium castaneum involves the activity of lipases, esterases, and laccase2.

    Science.gov (United States)

    Julio, Alison Henrique Ferreira; Gigliolli, Adriana Aparecida Sinópolis; Cardoso, Kátia Aparecida Kern; Drosdoski, Sandro Daniel; Kulza, Rodrigo Amaral; Seixas, Flávio Augusto Vicente; Ruvolo-Takasusuki, Maria Claudia Colla; de Souza, Cristina Giatti Marques; Lapenta, Ana Silvia

    2017-05-01

    Several recent studies have elucidated the molecular mechanisms that confer insecticide resistance on insect pests. However, little is known about multiple resistance in red flour beetle (Tribolium castaneum) at molecular level. The multiple resistance is characterized as resistance to different classes of insecticides that have different target sites, and is mediated by several enzymatic systems. In this study, we investigated the biochemical and molecular mechanisms involved in multiple resistance of T. castaneum to bifenthrin (pyrethroid [Pyr]) and pirimiphos-methyl (organophosphate [Org]). We used artificial selection, biochemical and in silico approaches including structural computational biology. After five generations of artificial selection in the presence of bifenthrin (F5Pyr) or pirimiphos-methyl (F5Org), we found high levels of multiple resistance. The hierarchical enzymatic cluster revealed a pool of esterases (E), lipases (LIPs) and laccase2 (LAC2) potentially contributing to the resistance in different ways throughout development, after one or more generations in the presence of insecticides. The enzyme-insecticide interaction network indicated that E2, E3, LIP3, and LAC2 are enzymes potentially required for multiple resistance phenotype. Kinetic analysis of esterases from F5Pyr and F5Org showed that pirimiphos-methyl and specially bifenthrin promote enzyme inhibition, indicating that esterases mediate resistance by sequestering bifenthrin and pirimiphos-methyl. Our computational data were in accordance with kinetic results, indicating that bifenthrin has higher affinity at the active site of esterase than pirimiphos-methyl. We also report the capability of these insecticides to modify the development in T. castaneum. Our study provide insights into the biochemical mechanisms employed by T. castaneum to acquire multiple resistance. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Biochemical Characterization and Relative Expression Levels of Multiple Carbohydrate Esterases of the Xylanolytic Rumen Bacterium Prevotella ruminicola 23 Grown on an Ester-Enriched Substrate ▿ †

    Science.gov (United States)

    Kabel, Mirjam A.; Yeoman, Carl J.; Han, Yejun; Dodd, Dylan; Abbas, Charles A.; de Bont, Jan A. M.; Morrison, Mark; Cann, Isaac K. O.; Mackie, Roderick I.

    2011-01-01

    We measured expression and used biochemical characterization of multiple carbohydrate esterases by the xylanolytic rumen bacterium Prevotella ruminicola 23 grown on an ester-enriched substrate to gain insight into the carbohydrate esterase activities of this hemicellulolytic rumen bacterium. The P. ruminicola 23 genome contains 16 genes predicted to encode carbohydrate esterase activity, and based on microarray data, four of these were upregulated >2-fold at the transcriptional level during growth on an ester-enriched oligosaccharide (XOSFA,Ac) from corn relative to a nonesterified fraction of corn oligosaccharides (AXOS). Four of the 16 esterases (Xyn10D-Fae1A, Axe1-6A, AxeA1, and Axe7A), including the two most highly induced esterases (Xyn10D-Fae1A and Axe1-6A), were heterologously expressed in Escherichia coli, purified, and biochemically characterized. All four enzymes showed the highest activity at physiologically relevant pH (6 to 7) and temperature (30 to 40°C) ranges. The P. ruminicola 23 Xyn10D-Fae1A (a carbohydrate esterase [CE] family 1 enzyme) released ferulic acid from methylferulate, wheat bran, corn fiber, and XOSFA,Ac, a corn fiber-derived substrate enriched in O-acetyl and ferulic acid esters, but exhibited negligible activity on sugar acetates. As expected, the P. ruminicola Axe1-6A enzyme, which was predicted to possess two distinct esterase family domains (CE1 and CE6), released ferulic acid from the same substrates as Xyn10D-Fae1 and was also able to cleave O-acetyl ester bonds from various acetylated oligosaccharides (AcXOS). The P. ruminicola 23 AxeA1, which is not assigned to a CE family, and Axe7A (CE7) were found to be acetyl esterases that had activity toward a broad range of mostly nonpolymeric acetylated substrates along with AcXOS. All enzymes were inhibited by the proximal location of other side groups like 4-O-methylglucuronic acid, ferulic acid, or acetyl groups. The unique diversity of carbohydrate esterases in P. ruminicola 23

  5. Crystallization and preliminary crystallographic studies of the metalloglycoprotein esterase A4 using a baculovirus expression system

    Energy Technology Data Exchange (ETDEWEB)

    Hiraki, Toshiki [Protein Design Laboratory, Yokohama City University, 1-7-29 Suehiro, Tsurumi, Yokohama 230-0045 (Japan); Shibayama, Naoya [Department of Physiology, Division of Biophysics, Jichi Medical University, 3311-1 Yakushiji, Shimotsuke, Tochigi 329-0498 (Japan); Yoon, Young-Ho [Protein Design Laboratory, Yokohama City University, 1-7-29 Suehiro, Tsurumi, Yokohama 230-0045 (Japan); Yun, Kyung-Mook [Department of Physiology, Division of Biophysics, Jichi Medical University, 3311-1 Yakushiji, Shimotsuke, Tochigi 329-0498 (Japan); Hamamoto, Toshiro [Department of Biochemistry, Jichi Medical University, 3311-1 Yakushiji, Shimotsuke, Tochigi 329-0498 (Japan); Tame, Jeremy R. H.; Park, Sam-Yong, E-mail: park@tsurumi.yokohama-cu.ac.jp [Protein Design Laboratory, Yokohama City University, 1-7-29 Suehiro, Tsurumi, Yokohama 230-0045 (Japan)

    2007-09-01

    Esterase A4 (EA4) is a timer protein found in diapause eggs of the silkworm Bombyx mori. The gene for this metalloglycoprotein was cloned from B. mori eggs and expressed using a baculovirus expression system in silkworm pupae. Crystals of the purified protein have been grown that diffract to beyond 2.1 Å resolution at 100 K using synchrotron radiation. Esterase A4 (EA4) is a timer protein found in diapause eggs of the silkworm Bombyx mori. The gene for this metalloglycoprotein was cloned from B. mori eggs and expressed using a baculovirus expression system in silkworm pupae. Crystals of the purified protein have been grown that diffract to beyond 2.1 Å resolution at 100 K using synchrotron radiation. The protein crystals belong to space group P2{sub 1}, with unit-cell parameters a = 47.1, b = 73.9, c = 47.4 Å, β = 104.1°. With one dimer per asymmetric unit, the crystal volume per unit protein weight (V{sub M}) is 2.3 Å{sup 3} Da{sup −1} and the solvent content is 47%.

  6. 21 CFR 173.140 - Esterase-lipase derived from Mucor miehei.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Esterase-lipase derived from Mucor miehei. 173.140... HUMAN CONSUMPTION Enzyme Preparations and Microorganisms § 173.140 Esterase-lipase derived from Mucor miehei. Esterase-lipase enzyme, consisting of enzyme derived from Mucor miehei var. Cooney et Emerson by...

  7. Esterase screening using whole cells of Brazilian soil microorganisms

    Energy Technology Data Exchange (ETDEWEB)

    Mantovani, Simone M.; Oliveira, Luciana G. de; Marsaioli, Anita J., E-mail: anita@iqm.unicamp.b [Universidade Estadual de Campinas (IQ/UNICAMP), SP (Brazil). Inst. de Quimica

    2010-07-01

    A miniaturized enzymatic assay using fluorescent probes to reveal esterase producing microorganisms was optimized and applied to screen 64 soil bacterial strains. The best results were validated using traditional non-fluorogenic assays with acetyl and propanoyl phenylethanol to confirm the miniaturized results. The most active microorganisms belong to the genus Bacillus showing esterase activity and good enantiomeric ratios for the resolution of phenylethanol derivatives (E > 30). Part of the microorganisms are kept in our laboratory in glycerol or freezedried and the best microorganisms will be deposited in the CBMAI/CPQBA/UNICAMP culture collection. (author)

  8. Identification of novel esterase-active enzymes from hot environments by use of the host bacterium Thermus thermophilus

    Directory of Open Access Journals (Sweden)

    Benedikt eLeis

    2015-04-01

    Full Text Available Functional metagenomic screening strategies, which are independent of known sequence information, can lead to the identification of truly novel genes and enzymes. Since E. coli has been used exhaustively for this purpose as a host, it is important to establish alternative expression hosts and to use them for functional metagenomic screening for new enzymes. In this study we show that Thermus thermophilus HB27 is an excellent screening host and can be used as an alternative provider of truly novel biocatalysts. In a previous study we constructed the mutant strain BL03 that was no longer able to grow on defined minimal medium supplemented with tributyrin as the sole carbon source and could be used as a host to screen for metagenomic DNA fragments that could complement growth on tributyrin. Several thousand single fosmid clones from thermophilic metagenomic libraries from heated compost and hot spring water samples were subjected to a comparative screening for esterase activity in both T. thermophilus strain BL03 and E. coli EPI300. We scored a greater number of active clones in the thermophilic bacterium than in the mesophilic E. coli. From all clones functionally screened in E. coli, only two thermostable α/β-fold hydrolase enzymes with high amino acid sequence similarity to already characterized enzymes were identifiable. In contrast, five further fosmids were found that conferred lipolytic activities in T. thermophilus. Four open reading frames (ORFs were found which did not share significant similarity to known esterase enzymes. Two of the genes were expressed in both hosts and the novel thermophilic esterases, which based on their primary structures could not be assigned to known esterase or lipase families, were purified and preliminarily characterized. Our work underscores the benefit of using additional screening hosts other than E. coli for the identification of novel biocatalysts with industrial relevance.

  9. E3Net: a system for exploring E3-mediated regulatory networks of cellular functions.

    Science.gov (United States)

    Han, Youngwoong; Lee, Hodong; Park, Jong C; Yi, Gwan-Su

    2012-04-01

    Ubiquitin-protein ligase (E3) is a key enzyme targeting specific substrates in diverse cellular processes for ubiquitination and degradation. The existing findings of substrate specificity of E3 are, however, scattered over a number of resources, making it difficult to study them together with an integrative view. Here we present E3Net, a web-based system that provides a comprehensive collection of available E3-substrate specificities and a systematic framework for the analysis of E3-mediated regulatory networks of diverse cellular functions. Currently, E3Net contains 2201 E3s and 4896 substrates in 427 organisms and 1671 E3-substrate specific relations between 493 E3s and 1277 substrates in 42 organisms, extracted mainly from MEDLINE abstracts and UniProt comments with an automatic text mining method and additional manual inspection and partly from high throughput experiment data and public ubiquitination databases. The significant functions and pathways of the extracted E3-specific substrate groups were identified from a functional enrichment analysis with 12 functional category resources for molecular functions, protein families, protein complexes, pathways, cellular processes, cellular localization, and diseases. E3Net includes interactive analysis and navigation tools that make it possible to build an integrative view of E3-substrate networks and their correlated functions with graphical illustrations and summarized descriptions. As a result, E3Net provides a comprehensive resource of E3s, substrates, and their functional implications summarized from the regulatory network structures of E3-specific substrate groups and their correlated functions. This resource will facilitate further in-depth investigation of ubiquitination-dependent regulatory mechanisms. E3Net is freely available online at http://pnet.kaist.ac.kr/e3net.

  10. E3 Travel & Mission Support System

    Data.gov (United States)

    US Agency for International Development — ETRAMS is a travel data collection system developed by the CKM team in E3 that collects information on both the basic details of an employee's trips (destination,...

  11. E3 Financing How-to Guide

    Science.gov (United States)

    The Financing How-to-Guide is intended to help manufacturers and their communities navigate financing and investment opportunities. While this guide provides an overview, there is no one-way to pay for E3 activities or attract investment.

  12. E3: Economy, Energy and Environment

    Science.gov (United States)

    E3 is a technical assistance framework helping communities, manufacturers, and manufacturing supply chains adapt and thrive in today's green economy. Find information on pollution prevention, sustainable business practices, and energy efficiency.

  13. E3: Extreme Energy Event monitoring

    CERN Document Server

    CERN. Geneva

    2015-01-01

    World peace through CERN technology: Use of explosives especially in urban areas is the defining phenomenon of the last and current century. E3 aims to reduce such excesses of violence by collecting scientific hard evidence.

  14. Activity of pectin methyl esterase during blanching of peaches

    NARCIS (Netherlands)

    Tijskens, L.M.M.; Rodis, P.S.; Hertog, M.L.A.T.M.; Proxenia, N.; Dijk, van C.

    1999-01-01

    The activity of pectin methyl esterase (PE) in peaches during blanching treatments was modelled and analyzed. It was postulated that the enzyme exists in two configurations, one bound and one soluble. The bound configuration can be converted into the soluble configuration. These two configurations

  15. New cholesterol esterase inhibitors based on rhodanine and thiazolidinedione scaffolds

    DEFF Research Database (Denmark)

    Heng, Sabrina; Tieu, William; Hautmann, Stephanie

    2011-01-01

    We present a new class of inhibitors of pancreatic cholesterol esterase (CEase) based on 'priviledged' 5-benzylidenerhodanine and 5-benzylidene-2,4-thiazolidinedione structural scaffolds. The lead structures (5-benzylidenerhodanine 4a and 5-benzylidene-2,4-thiazolidinedione 4b) were identified in...

  16. The Lp_3561 and Lp_3562 enzymes support a functional divergence process in the lipase/esterase toolkit from Lactobacillus plantarum

    Directory of Open Access Journals (Sweden)

    Maria Esteban-Torres

    2016-07-01

    Full Text Available Lactobacillus plantarum species is a good source of esterases since both lipolytic and esterase activities have been described for strains of this species. No fundamental biochemical difference exists among esterases and lipases since both share a common catalytic mechanism. L. plantarum WCFS1 possesses a protein, Lp_3561, which is 44% identical to a previously described lipase, Lp_3562. In contrast to Lp_3562, Lp_3561 was unable to degrade esters possessing a chain length higher than C4 and the triglyceride tributyrin. As in other L. plantarum esterases, the electrostatic potential surface around the active site in Lp_3561 is predicted to be basic, whereas it is essentially neutral in the Lp_3562 lipase. The fact that the genes encoding both proteins were located contiguously in the L. plantarum WCFS1 genome, suggests that they originated by tandem duplication, and therefore are paralogs as new functions have arisen during evolution. The presence of the contiguous lp_3561 and lp_3562 genes was studied among L. plantarum strains. They are located in a 8,903 bp DNA fragment that encodes proteins involved in the catabolism of sialic acid and are predicted to increase bacterial adaptability under certain growth conditions.

  17. Esterase polymorphism marking cultivars of Manihot esculenta, Crantz

    Directory of Open Access Journals (Sweden)

    Adriana Gazoli Resende

    2004-07-01

    Full Text Available Esterase isozymes were used to detected substrate-preference polymorphism in twenty cultivars of Manihot esculenta, and to show cultivar-specific variation of this species. A relatively complex extraction solution of proteins from leaves was needed to show a larger number of esterase isozymes. Similarity between cultivars from six groups ranged from 51 to 96%. The cultivars identified by the same name seemed to be biochemically different regarding esterase isozymes. Esterase isozyme electrophoretic patterns could, therefore, be used to discriminate the cultivars identified by the same name, and to monitor the vegetative propagation of cultivars maintained in the germplasm collection. In breeding strategies, isoesterase analysis could be used to avoid intercrossing between the similar genotypes.Isoenzimas esterases foram usadas no presente estudo, para detectar polimorfismos específicos para diferentes substratos em vinte cultivares de Manihot esculenta, e para mostrar variações específicas de cultivares nesta espécie. Os diferentes cultivares de M. esculenta tem sido mantidos na coleção de germoplasma do Departamento de Agronomia da Universidade Estadual de Maringá (Maringá, PR, e foram provenientes de cultivares tradicionais coletados nas regiões sudoeste e noroeste do Estado. Foi necessário a utilização de uma solução de extração de proteínas relativamente mais complexa, para evidenciar um maior número de isoenzimas esterases. A similaridade entre os cultivares variou de 51 a 96%. Cultivares identificados pelo mesmo nome parecem ser bioquimicamente diferentes para as isoenzimas esterases. Os padrões eletroforéticos das isoesterases podem, portanto, serem usados para discriminar os cultivares que são identificados pelo mesmo nome, e para monitorar a propagação vegetativa dos cultivares mantidos na coleção de germoplasma. A análise das isoesterases pode também ser usada para evitar cruzamentos entre genótipos mais

  18. Esterase activity able to hydrolyze dietary antioxidant hydroxycinnamates is distributed along the intestine of mammals

    DEFF Research Database (Denmark)

    Andreasen, Mette Findal; Kroon, P A; Williamson, G

    2001-01-01

    and may contribute to the beneficial effects derived from consumption of cereal bran. However, these compounds are ester linked to the main polymers in the plant cell wall and cannot be absorbed in this complex form. The present work shows that esterases with activity toward esters of the major dietary...... hydroxycinnamates are distributed throughout the intestinal tract of mammals. In rats, the cinnamoyl esterase activity in the small intestine is derived mainly from the mucosa, whereas in the large intestine the esterase activity was found predominantly in the luminal microflora. Mucosa cell-free extracts obtained...... from human duodenum, jejunum, and ileum efficiently hydrolyzed various hydroxycinnamoyl esters, providing the first evidence of human cinnamoyl esterase(s). This study first demonstrates the release by human colonic esterase(s) (mostly of microbial origin) of sinapic acid and p-coumaric acid from rye...

  19. In vitro comparison of rat and chicken brain neurotoxic esterase

    International Nuclear Information System (INIS)

    Novak, R.; Padilla, S.

    1986-01-01

    A systematic comparison was undertaken to characterize neurotoxic esterase (NTE) from rat and chicken brain in terms of inhibitor sensitivities, pH optima, and molecular weights. Paraoxon titration of phenyl valerate (PV)-hydrolyzing carboxylesterases showed that rat esterases were more sensitive than chicken to paraoxon inhibition at concentrations less than or equal to microM and superimposable with chicken esterases at concentrations of 2.5-1000 microM. Mipafox titration of the paraoxon-resistant esterases at a fixed paraoxon concentration of 100 microM (mipafox concentration: 0-1000 microM) resulted in a mipafox I50 of 7.3 microM for chicken brain NTE and 11.6 microM for rat brain NTE. NTE (i.e., paraoxon-resistant, mipafox-sensitive esterase activity) comprised 80% of chicken and 60% of rat brain paraoxon-resistant activity with the specific activity of chicken brain NTE approximately twice that of rat brain NTE. The pH maxima for NTE from both species was similar showing broad, slightly alkaline optima from pH 7.9 to 8.6. [ 3 H]Diisopropyl phosphorofluoridate (DFP)-labeled NTE from the brains of both species had an apparent mol wt of 160,000 measured by sodium dodecyl sulfate polyacrylamide gel electrophoresis. In conclusion, NTE from both species was very similar, with the mipafox I50 for rat NTE within the range of reported values for chicken and human NTE, and the inhibitor parameters of the chicken NTE assay were applicable for the rat NTE assay

  20. Analysis of esterase isozyme and SSR for mutagenic progenies induced by space mutation in mustard

    International Nuclear Information System (INIS)

    Shen Jinjuan; Liu Yihua; Zhang Zhaorong; Ran Guangkui; Zhao Shouzhong; Xiao Li

    2012-01-01

    Seeds of five mustard (Brassica juncea Coss) varieties were carried into outer space by 'Shijian No.8' satellite. After five years' consecutive planting and selection, ten relatively stable mutant lines were obtained, which had significant variation in agronomic and economic characters. The mutant lines and their original varieties without space mutation treatment as control were studied by esterase isozyme and SSR analyses. Electrophoresis analysis of esterase isozymes indicated that there were differences between mutant lines and their controls in enzyme types and enzyme activity. Different mustard varieties had different enzymographs, and so did the mutants induced by space mutation, which shows different sensitivity among different mustard varieties. The SSR analysis showed that large differences were found in the SSR loci between mutant lines and their original variety, the variation frequency was between 9.52% and 57.14% with an average frequency of 26.19% for all the mutant lines. Among the mutant SSR loci, about 56.36% showed changes in band number and 43.64% in molecular weight. These results indicated that the ten mutant lines had large genetic difference in phenotype, genomic sequence and gene expression, and the outer space mutation would be an effective method to develop new mustard germplasm and variety. (authors)

  1. The monotony of transferrin and esterase electrophoretic patterns in pirarucu, Arapaima gigas (Schinz, 1822) from Santa Cruz Lake, Tefé River, Amazonas, Brazil.

    Science.gov (United States)

    Teixeira, A S

    2008-05-07

    Starch gel electrophoresis was used for examining the transferrin gene locus (Tf) and two esterase gene loci (Est-1 and Est-D1) of a pirarucu (Arapaima gigas) population sample collected from Santa Cruz Lake, Tefé River, Amazonas, Brazil. The Tf locus was tentatively classified as being polymorphic, showing two double-banded patterns (Tf(12) and Tf(22)) of the three theoretically expected ones (Tf(11), Tf(12) and Tf(22)), presumably controlled by two co-dominant alleles, Tf(1) and Tf(2). The monotony detected in pirarucu Tf locus genotypes showing a very high proportion of the double-banded heterozygote pattern Tf(12) (95% of the sampled individuals) may indicate the possibility of their having come from representatives of the same brood begotten by a pair of fish, where a single-banded Tf(11) homozygote pattern male would have crossed with a single-banded Tf(22) homozygote pattern female, or vice versa. One zone of electrophoretic activity was detected in esterase, presumably controlled by a monomorphic Est-1 locus with the fixed allele Est-1(1) where all individuals showed the single-banded Est-1(11) homozygote pattern. Esterase-D also displayed one zone of electrophoretic activity, presumably controlled by a monomorphic Est-D1 locus with a fixed allele Est-D1(1) where all individuals revealed the single-banded Est-D1(11) genotype pattern. The monotony comprised by single-banded genotype patterns in both esterase systems tested may also indicate the possibility of the individuals from the sample examined having come from representatives of the same brood begotten by a pair of fish with both the male and female having the same genotypes.

  2. Leucocyte esterase in the rapid diagnosis of paediatric septic arthritis.

    LENUS (Irish Health Repository)

    Kelly, E G

    2013-02-01

    Septic arthritis may affect any age group but is more common in the paediatric population. Infection is generally bacterial in nature. Prompt diagnosis is crucial, as delayed treatment is associated with lifelong joint dysfunction. A clinical history and application of Kocher\\'s criteria may indicate that there is a septic arthritis. However, definitive diagnosis is made on culture of septic synovial fluid. The culture process can take over 24h for the initial culture to yield bacterial colonies. Leucocyte esterase is released by leucocytes at the site of an infection. We hypothesise that leucocyte esterase can be utilized in the rapid diagnosis of septic arthritis and shorten the time to decisive treatment whilst simultaneously decreasing unnecessary treatment of non-septic joints.

  3. Inhibition of Pectin Methyl Esterase Activity By Green Tea Catechins

    OpenAIRE

    Sagi, Irit; Lewis, Kristin; Tworowski, Dmitry; Shahar, Chen; Selzer, Tzvia

    2008-01-01

    Pectin methyl esterases (PMEs) and their endogenous inhibitors are involved in the regulation of many processes in plant physiology, ranging from tissue growth and fruit ripening to parasitic plant haustorial formation and host invasion. Thus, control of PME activity is critical for enhancing our understanding of plant physiological processes and regulation. Here we report on the identification of epigallocatechin gallate (EGCG), a green tea component, as a natural inhibitor for pectin ...

  4. 3 Benzyl-6-chloropyrone: a suicide inhibitor of cholesterol esterase

    International Nuclear Information System (INIS)

    Saint, C.; Gallo, I.; Kantorow, M.; Bailey, J.M.

    1986-01-01

    Cholesterol, absorbed from the intestine, appears in lymph as the ester. Cholesterol esterase is essential for this process, since depletion of the enzyme blocks and repletion restores, absorption. Selective inhibitors of cholesterol esterase may thus prove useful in reducing cholesterol uptake. A series of potential suicide substrates were synthesized which, following cleavage by the enzyme, would attack the putative nucleophile in the active site. One of these, 3-benzyl-6-chloropyrone (3BCP), inhibited both synthesis and hydrolysis of 14 C-cholesteryl oleate with an I 50 of approximately 150 μM. The inactivation was time-dependent and characteristic of a suicide mechanism. The α pyrone structure (lactone analog) is cleaved by a serine-hydroxyl in the active site. This generates an enoyl chloride which inactivates the imidazole believed to play a part in the catalytic function of the enzyme. Inhibition by 3BCP is selective for cholesterol esterase. The activity of pancreatic lipase as not affected by concentrations up to 1 mM

  5. Inhibition of pectin methyl esterase activity by green tea catechins.

    Science.gov (United States)

    Lewis, Kristin C; Selzer, Tzvia; Shahar, Chen; Udi, Yael; Tworowski, Dmitry; Sagi, Irit

    2008-10-01

    Pectin methyl esterases (PMEs) and their endogenous inhibitors are involved in the regulation of many processes in plant physiology, ranging from tissue growth and fruit ripening to parasitic plant haustorial formation and host invasion. Thus, control of PME activity is critical for enhancing our understanding of plant physiological processes and regulation. Here, we report on the identification of epigallocatechin gallate (EGCG), a green tea component, as a natural inhibitor for pectin methyl esterases. In a gel assay for PME activity, EGCG blocked esterase activity of pure PME as well as PME extracts from citrus and from parasitic plants. Fluorometric tests were used to determine the IC50 for a synthetic substrate. Molecular docking analysis of PME and EGCG suggests close interaction of EGCG with the catalytic cleft of PME. Inhibition of PME by the green tea compound, EGCG, provides the means to study the diverse roles of PMEs in cell wall metabolism and plant development. In addition, this study introduces the use of EGCG as natural product to be used in the food industry and agriculture.

  6. A comparison between activities for non-specific esterases and esterproteases

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D

    1988-01-01

    Electrophoretic separation of non-specific esterases and esterproteases from kidney, lung, and liver have been carried out in polyacrylamide gels. By use of zone electrophoresis, isoelectric focusing, and 2-dimensional electrophoresis it was found that most of the esterprotease bands had the same...... localization in the gels as non-specific esterase bands. A number of esterase bands showed no activity towards the esterprotease substrates and a single kidney band possessed esterprotease activity only. Isozymes of the ES-6 and ES-9 zones showed sex dependent esterprotease reactions. In sections esterase...

  7. Characterisation of a New Family of Carboxyl Esterases with an OsmC Domain.

    Directory of Open Access Journals (Sweden)

    Mai-Britt V Jensen

    Full Text Available Proteins in the serine esterase family are widely distributed in bacterial phyla and display activity against a range of biologically produced and chemically synthesized esters. A serine esterase from the psychrophilic bacterium Pseudoalteromonas arctica with a C-terminal OsmC-like domain was recently characterized; here we report on the identification and characterization of further putative esterases with OsmC-like domains constituting a new esterase family that is found in a variety of bacterial species from different environmental niches. All of these proteins contained the Ser-Asp-His motif common to serine esterases and a highly conserved pentapeptide nucleophilic elbow motif. We produced these proteins heterologously in Escherichia coli and demonstrated their activity against a range of esterase substrates. Two of the esterases characterized have activity of over two orders of magnitude higher than other members of the family, and are active over a wide temperature range. We determined the crystal structure of the esterase domain of the protein from Rhodothermus marinus and show that it conforms to the classical α/β hydrolase fold with an extended 'lid' region, which occludes the active site of the protein in the crystal. The expansion of characterized members of the esterase family and demonstration of activity over a wide-range of temperatures could be of use in biotechnological applications such as the pharmaceutical, detergent, bioremediation and dairy industries.

  8. A new microplate screening method for the simultaneous activity quantification of feruloyl esterases, tannases, and chlorogenate esterases.

    Science.gov (United States)

    Ramírez, L; Arrizon, J; Sandoval, G; Cardador, A; Bello-Mendoza, R; Lappe, P; Mateos-Díaz, J C

    2008-12-01

    Feruloyl, chlorogenate esterases, and tannases are enzymes useful in phenolic modifications of pharmaceutical relevance as protectors against several degenerative human diseases. Therefore, there is a growing interest in discovering new sources of these enzymes. However, traditional methods for their activity measurements are time-consuming and poorly adapted for high-throughput screening. In this study, a successful new microplate high-throughput screening method for the simultaneous quantification of all mentioned activities is demonstrated. This method allows the detection of activities as low as 1.7 mU ml(-1). Furthermore, reaction rates increased proportionally with the amount of enzyme added, and no interferences with the other commercial hydrolases tested were found. The utility of the method was demonstrated after simultaneously screening feruloyl, chlorogenate esterase, and tannase activities in solid state fermentation extracts obtained during the kinetics of production of 20 fungal strains. Among these, seven strains were positive for at least one of the esterase activities tested. This result shows the potential for the rapid routine screening assays for multiple samples of moderate low to high enzymatic levels.

  9. Extracellular esterases of phylloplane yeast Pseudozyma antarctica induce defect on cuticle layer structure and water-holding ability of plant leaves.

    Science.gov (United States)

    Ueda, Hirokazu; Mitsuhara, Ichiro; Tabata, Jun; Kugimiya, Soichi; Watanabe, Takashi; Suzuki, Ken; Yoshida, Shigenobu; Kitamoto, Hiroko

    2015-08-01

    Aerial plant surface (phylloplane) is a primary key habitat for many microorganisms but is generally recognized as limited in nutrient resources. Pseudozyma antarctica, a nonpathogenic yeast, is commonly isolated from plant surfaces and characterized as an esterase producer with fatty acid assimilation ability. In order to elucidate the biological functions of these esterases, culture filtrate with high esterase activity (crude enzyme) of P. antarctica was applied onto leaves of tomato and Arabidopsis. These leaves showed a wilty phenotype, which is typically associated with water deficiency. Furthermore, we confirmed that crude enzyme-treated detached leaves clearly lost their water-holding ability. In treated leaves of both plants, genes associated to abscisic acid (ABA; a plant stress hormone responding osmotic stress) were activated and accumulation of ABA was confirmed in tomato plants. Microscopic observation of treated leaf surfaces revealed that cuticle layer covering the aerial epidermis of leaves became thinner. A gas chromatography-mass spectrometry (GC-MS) analysis exhibited that fatty acids with 16 and 18 carbon chains were released in larger amounts from treated leaf surfaces, indicating that the crude enzyme has ability to degrade lipid components of cuticle layer. Among the three esterases detected in the crude enzyme, lipase A, lipase B, and P. antarctica esterase (PaE), an in vitro enzyme assay using para-nitrophenyl palmitate as substrate demonstrated that PaE was the most responsible for the degradation. These results suggest that PaE has a potential role in the extraction of fatty acids from plant surfaces, making them available for the growth of phylloplane yeasts.

  10. Scaling laws in (e,3e) processes

    International Nuclear Information System (INIS)

    Gasaneo, G; Rodriguez, K V; Ancarani, L U; Cappello, C Dal; Charpentier, I

    2009-01-01

    We study the double ionization of helium-like ions by impact of electrons with high incident energy. Within the isoelectronic sequence, an approximate scaling law for (e,3e) differential cross sections is proposed and confirmed by calculations. The latter are performed using 14-parameters Hylleraas-like wave functions to represent the bound electrons in the initial channel, plane waves for the fast incoming and scattered electrons, and a continuum distorted wave approach for the two ejected electrons in the final channel.

  11. E3 Success Story - E3 Southwest Virginia: Economy, Energy and the Environment

    Science.gov (United States)

    E3 Southwest Virginia supports sustainable manufacturing in 17 counties in southwest Virginia. The MTC provides manufacturers with assessments of production processes to reduce their energy consumption and drive innovation.

  12. Alicyclobacillus acidocaldarius Thermophilic Esterase EST2's Activity in Milk and Cheese Models

    NARCIS (Netherlands)

    Mandrich, L.; Manco, M.; Rossie, M.; Floris, E.; Jansen-van den Bosch, T.; Smit, G.; Wouters, J.A.

    2006-01-01

    The aim of this work was to investigate the behavior of thermophilic esterase EST2 from Alicyclobacillus acidocaldarius in milk and cheese models. The pure enzyme was used to compare the EST2 hydrolytic activity to the activity of endogenous esterase EstA from Lactococcus lactis. The results

  13. Functional and structural characterization of a thermostable acetyl esterase from Thermotoga maritima

    NARCIS (Netherlands)

    Levisson, M.; Han, G.W.; Deller, M.C.; Hendriks, S.N.A.; Oost, van der J.; Kengen, S.W.M.

    2012-01-01

    TM0077 from Thermotoga maritima is a member of the carbohydrate esterase family 7 and is active on a variety of acetylated compounds, including cephalosporin C. TM0077 esterase activity is confined to short-chain acyl esters (C2-C3), and is optimal around 100°C and pH 7.5. The positional specificity

  14. Esterases in striated muscle from mice with the Chediak-Higashi syndrome

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D

    1981-01-01

    In this paper a localized strong reaction for non-specific esterase forming cylindric structures is described within skeletal muscle fibres from the beige mouse. It seems from zymograms and protein electrophoresis that this esterase is membrane bound, highly reactive and present in rather small...

  15. Esterase Isoenzyme Profiles in Acute and Chronic Leukemias.

    Science.gov (United States)

    Drexler, H G; Gignac, S M; Hoffbrand, A V; Minowada, J

    1991-01-01

    Using isoelectric focusing (IEF) a number of carboxylic esterase isoenzymes (EC 3.1.1.1) with isoelectric points between pH 4.5-8.0 can be separated. One particular isoenzyme with an isoelectric point at about pH 6.0, the Mono-band, can be selectively and completely inhibited by sodium fluoride; this isoenzyme comprises a number of closely related subcomponents and may appear in more than one band on the gel. We analyzed the expression of typical esterase isoenzyme patterns in cells from a large panel of leukemias which were tested under identical conditions by IEF on horizontal thin-layer polyacrylamide gels with an ampholyte of pH 2-11. The 442 cases of acute and chronic myeloid and lymphoid leukemia (AML/AMMoL, CML/CMML, ALL, CLL) were classified according to clinical, morpho-cytochemical and immunophenotyping criteria. While bands between pH 4.5-5.5 appeared not to be specific for lineage or stage of differentiation, isoenzymes between pH 6.6-7.7 provided information on the type of leukemia involved. Seven typical isoenzyme patterns termed Mono1/Mono2 (fo monocyte-associated), My1/My2 (myeloid), Lym1/Lym2 (lymphoid) and Und (undifferentiated) could be discerned. Lym and Und patterns are characterized by fewer bands with a weaker staining intensity than Mono and My patterns. Nearly all cases of lymphoid leukemias (acute and chronic) expressed only Lym or Und esterase isoenzyme patterns, but no Mono or My patterns. Cases of acute or chronic myeloid and (myelo)monocytic leukemia showed strong isoenzyme staining displaying predominantly Mono or My isoenzyme patterns. The isoenzyme patterns found in CML in lymphoid or myeloid blast crisis corresponded to those seen in the respective acute leukemias, ALL or AML. The Mono-band was found in most cases of leukemias with monocytic elements (AMMoL 80%, CML 44%, CMML 100%), in the occasional case of CML-myeloid blast crisis or AML, but in none of the cases of ALL or CLL. This isoenzyme is a distinctive, specific marker for

  16. Feruloyl esterase from Aspergillus clavatus improves xylan hydrolysis of sugarcane bagasse

    Directory of Open Access Journals (Sweden)

    Dyoni M. de Oliveira

    2016-12-01

    Full Text Available Feruloyl esterase is a subclass of carboxylic acid esterases with the capacity to release ferulic acid and other cinnamic acids from plant cell walls and synthetic substrates. Feruloyl esterases act synergistically with xylanases removing ferulic acid residues esterified to arabinoxylans. Feruloyl esterase type D from Aspergillus clavatus (AcFAE was expressed in Escherichia coli, purified, and applied with a commercial xylanase consortium (Novozymes for hydrolysis of sugarcane bagasse. Feruloyl esterase plus xylanase increased 5.13-fold the releasing of ferulic acid from sugarcane bagasse. Removal of only 7.7% of ferulic acid content by AcFAE increased 97.3% the sugarcane bagasse hydrolysis by xylanase. These data support the use of AcFAE as an interesting adjuvant enzyme to improve lignocellulose digestion and biotechnological tool for biorefineries.

  17. Organophosphate acetylcholine esterase inhibitor poisoning from a home-made shampoo.

    Science.gov (United States)

    Sadaka, Yair; Broides, Arnon; Tzion, Raffi Lev; Lifshitz, Matitiahu

    2011-07-01

    Organophosphate acetylcholine esterase inhibitor poisoning is a major health problem in children. We report an unusual cause of organophosphate acetylcholine esterase inhibitor poisoning. Two children were admitted to the pediatric intensive care unit due to organophosphate acetylcholine esterase inhibitor poisoning after exposure from a home-made shampoo that was used for the treatment of head lice. Owing to no obvious source of poisoning, the diagnosis of organophosphate acetylcholine esterase inhibitor poisoning in one of these patients was delayed. Both patients had an uneventful recovery. Organophosphate acetylcholine esterase inhibitor poisoning from home-made shampoo is possible. In cases where the mode of poisoning is unclear, direct questioning about the use of home-made shampoo is warranted, in these cases the skin and particularly the scalp should be rinsed thoroughly as soon as possible.

  18. Studies on the oxidizing system in Holt's medium for histochemical demonstration of esterase activity

    DEFF Research Database (Denmark)

    Kirkeby, S; Blecher, S R

    1978-01-01

    Esterase activity in guinea-pig thyroid and mouse epididymis epithelial cells has been studied using 5-bromoindoxyl acetate as substrate. The pattern of esterase activity in the thyroid of the guinea-pig is constant, irrespective of whether ferri-ferrocyanide (FFC) or certain copper compounds...... cells contain an esterase activity which is not inhibited by conventional SH blocking agents, nor by high concentrations of FFC. From these results it appears that the mode of action of FFC in Holt's medium is as follows. At low concentrations FFC appears to act primarily as a catalytic agent...... in oxidation of indoxyl to indigoid. At high concentration FFC acts as an inhibitor of guinea-pig thyroid esterase, by oxidation of SH groups in the active centre. The esterase of mouse epididymis cell type EH 1 is not subject to this inhibition by FFC, presumably because it does not contain accessible SH...

  19. C1-esterase inhibitor protects against early vein graft remodeling under arterial blood pressure.

    Science.gov (United States)

    Krijnen, Paul A J; Kupreishvili, Koba; de Vries, Margreet R; Schepers, Abbey; Stooker, Wim; Vonk, Alexander B A; Eijsman, Leon; Van Hinsbergh, Victor W M; Zeerleder, Sacha; Wouters, Diana; van Ham, Marieke; Quax, Paul H A; Niessen, Hans W M

    2012-01-01

    Arterial pressure induced vein graft injury can result in endothelial loss, accelerated atherosclerosis and vein graft failure. Inflammation, including complement activation, is assumed to play a pivotal role herein. Here, we analyzed the effects of C1-esterase inhibitor (C1inh) on early vein graft remodeling. Human saphenous vein graft segments (n=8) were perfused in vitro with autologous blood either supplemented or not with purified human C1inh at arterial pressure for 6h. The vein segments and perfusion blood were analyzed for cell damage and complement activation. In addition, the effect of purified C1inh on vein graft remodeling was analyzed in vivo in atherosclerotic C57Bl6/ApoE3 Leiden mice, wherein donor caval veins were interpositioned in the common carotid artery. Application of C1inh in the in vitro perfusion model resulted in significantly higher blood levels and significantly more depositions of C1inh in the vein wall. This coincided with a significant reduction in endothelial loss and deposition of C3d and C4d in the vein wall, especially in the circular layer, compared to vein segments perfused without supplemented C1inh. Administration of purified C1inh significantly inhibited vein graft intimal thickening in vivo in atherosclerotic C57Bl6/ApoE3 Leiden mice, wherein donor caval veins were interpositioned in the common carotid artery. C1inh significantly protects against early vein graft remodeling, including loss of endothelium and intimal thickening. These data suggest that it may be worth considering its use in patients undergoing coronary artery bypass grafting. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  20. Profiling and functional classification of esterases in olive (Olea europaea) pollen during germination.

    Science.gov (United States)

    Rejón, Juan D; Zienkiewicz, Agnieszka; Rodríguez-García, María Isabel; Castro, Antonio J

    2012-10-01

    A pollen grain contains a number of esterases, many of which are released upon contact with the stigma surface. However, the identity and function of most of these esterases remain unknown. In this work, esterases from olive pollen during its germination were identifided and functionally characterized. The esterolytic capacity of olive (Olea europaea) pollen was examined using in vitro and in-gel enzymatic assays with different enzyme substrates. The functional analysis of pollen esterases was achieved by inhibition assays by using specific inhibitors. The cellular localization of esterase activities was performed using histochemical methods. Olive pollen showed high levels of non-specific esterase activity, which remained steady after hydration and germination. Up to 20 esterolytic bands were identified on polyacrylamide gels. All the inhibitors decreased pollen germinability, but only diisopropyl fluorophosphate (DIFP) hampered pollen tube growth. Non-specific esterase activity is localized on the surface of oil bodies (OBs) and small vesicles, in the pollen intine and in the callose layer of the pollen tube wall. Acetylcholinesterase (AChE) activity was mostly observed in the apertures, exine and pollen coat, and attached to the pollen tube wall surface and to small cytoplasmic vesicles. In this work, for the first time a systematic functional characterization of esterase enzymes in pollen from a plant species with wet stigma has been carried out. Olive pollen esterases belong to four different functional groups: carboxylesterases, acetylesterases, AChEs and lipases. The cellular localization of esterase activity indicates that the intine is a putative storage site for esterolytic enzymes in olive pollen. Based on inhibition assays and cellular localization of enzymatic activities, it can be concluded that these enzymes are likely to be involved in pollen germination, and pollen tube growth and penetration of the stigma.

  1. Branched nanotrees with immobilized acetylcholine esterase for nanobiosensor applications

    Energy Technology Data Exchange (ETDEWEB)

    Risveden, Klas; Bhand, Sunil; Danielsson, Bengt [Department of Pure and Applied Biochemistry, Center for Chemistry and Chemical Engineering, Lund University, PO Box 124, SE-22100 Lund (Sweden); Dick, Kimberly A; Samuelson, Lars [Solid State Physics, Lund University, Box 118, S-22100 Lund (Sweden); Rydberg, Patrik, E-mail: Kimberly.Dick@ftf.lth.se [Department of Medicinal Chemistry, Faculty of Pharmaceutical Sciences, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen (Denmark)

    2010-02-05

    A novel lab-on-a-chip nanotree enzyme reactor is demonstrated for the detection of acetylcholine. The reactors are intended for use in the RISFET (regional ion sensitive field effect transistor) nanosensor, and are constructed from gold-tipped branched nanorod structures grown on SiN{sub x}-covered wafers. Two different reactors are shown: one with simple, one-dimensional nanorods and one with branched nanorod structures (nanotrees). Significantly higher enzymatic activity is found for the nanotree reactors than for the nanorod reactors, most likely due to the increased gold surface area and thereby higher enzyme binding capacity. A theoretical calculation is included to show how the enzyme kinetics and hence the sensitivity can be influenced and increased by the control of electrical fields in relation to the active sites of enzymes in an electronic biosensor. The possible effects of electrical fields employed in the RISFET on the function of acetylcholine esterase is investigated using quantum chemical methods, which show that the small electric field strengths used are unlikely to affect enzyme kinetics. Acetylcholine esterase activity is determined using choline oxidase and peroxidase by measuring the amount of choline formed using the chemiluminescent luminol reaction.

  2. Heterologous expression of two Aspergillus niger feruloyl esterases in Trichoderma reesei for the production of ferulic acid from wheat bran.

    Science.gov (United States)

    Long, Liangkun; Zhao, Haoyuan; Ding, Dafan; Xu, Meijuan; Ding, Shaojun

    2018-05-01

    Feruloyl esterase (FAE)-encoding genes AnfaeA and AnfaeB were isolated from Aspergillus niger 0913. For overexpression of the two genes in Trichoderma reesei, constitutive and inductive expression plasmids were constructed based on parental plasmid pAg1-H3. The constructed plasmids contained AnfaeA or AnfaeB gene under the control of glyceraldehyde-3-phosphate dehydrogenase A gene (gpdA) promoter (from A. nidulans) or cellobiohydrolases I (cbh I) gene promoter (from T. reesei), and cbh I terminator from T. reesei. The target plasmids were transferred into T. reesei D-86271 (Rut-C30) by Agrobacterium tumefaciens-mediated transformation (ATMT), respectively. A high level of feruloyl esterase was produced by the recombinant fungal strains under solid-state fermentation, and the cbh I promoter was more efficient than the gpdA promoter in the expression of AnfaeA. The optimum temperatures and pH values were 50 °C and 5.0 for AnFAEA, and 35 °C and 6.0 for AnFAEB. The maximum production levels were 20.69 U/gsd for AnFAEA and 15.08 U/gsd for AnFAEB. The recombinant fungal enzyme systems could release 62.9% (for AnFAEA) and 52.2% (for AnFAEB) of total ferulic acids from de-starched wheat bran, which was higher than the 46.3% releasing efficiency of A. niger 0913. The supplement of xylanase from T. longibrachiatum in the enzymatic hydrolysis led to a small increment of the ferulic acids release.

  3. E3 Success Story - San Antonio E3 Case Study: UEMC “Now Hiring”

    Science.gov (United States)

    exas Manufacturing Assistance Center conducted an E3 assessment as part of the local Lean-Clean-Energy program in October of 2009. Overall, UEMC has reduced demand by approximately 30kw and reduced consumption by over 100,000 kwh/year.

  4. Estrogenic and esterase-inhibiting potency in rainwater in relation to pesticide concentrations, sampling season and location

    International Nuclear Information System (INIS)

    Hamers, T.; Brink, P.J. van den; Mos, L.; Linden, S.C. van der; Legler, J.; Koeman, J.H.; Murk, A.J.

    2003-01-01

    Estrogenic potency of rainwater correlated well with organochlorine concentrations, but could not be attributed to specific pesticides. - In a year-round monitoring program (1998), pesticide composition and toxic potency of the mix of pollutants present in rainwater were measured. The goal of the study was to relate atmospheric deposition of toxic potency and pesticide composition to each other and to sampling period and local agricultural activity. Rainwater was collected in 26 consecutive periods of 14 days in a background location (BACK) and in two locations representative for different agricultural practices, i.e. intensive greenhouse horticulture (HORT) and flower bulb culture (BULB). Samples were chemically analyzed for carbamate (CARB), organophosphate (OP) and organochlorine (OC) pesticides and metabolites. Esterase inhibiting potency of rainwater extracts was measured in a specially developed bio-assay with honeybee esterases and was expressed as an equivalent concentration of the model inhibitor dichlorvos. Estrogenic potency of the extracts was measured in the ER-CALUX reporter gene assay and was expressed as an equivalent concentration of estradiol. Multivariate principal component analysis (PCA) techniques proved to be valuable tools to analyze the numerous pesticide concentrations in relation to toxic potency, sampling location, and sampling season. Pesticide composition in rainwater depended much more on sampling season than on sampling location, but differences between SPRING and SUMMER were mainly attributed to local differences in agricultural practice. On average, the esterase inhibiting potency exceeded the maximum permissible concentration set for dichlorvos in The Netherlands, and was significantly higher in HORT than in BACK and BULB. Esterase inhibition correlated significantly with OP and CARB concentrations, as expected given the working mechanism of these insecticides. The estrogenic potency incidentally exceeded NOEC levels reported for

  5. Estrogenic and esterase-inhibiting potency in rainwater in relation to pesticide concentrations, sampling season and location

    Energy Technology Data Exchange (ETDEWEB)

    Hamers, T.; Brink, P.J. van den; Mos, L.; Linden, S.C. van der; Legler, J.; Koeman, J.H.; Murk, A.J

    2003-05-01

    Estrogenic potency of rainwater correlated well with organochlorine concentrations, but could not be attributed to specific pesticides. - In a year-round monitoring program (1998), pesticide composition and toxic potency of the mix of pollutants present in rainwater were measured. The goal of the study was to relate atmospheric deposition of toxic potency and pesticide composition to each other and to sampling period and local agricultural activity. Rainwater was collected in 26 consecutive periods of 14 days in a background location (BACK) and in two locations representative for different agricultural practices, i.e. intensive greenhouse horticulture (HORT) and flower bulb culture (BULB). Samples were chemically analyzed for carbamate (CARB), organophosphate (OP) and organochlorine (OC) pesticides and metabolites. Esterase inhibiting potency of rainwater extracts was measured in a specially developed bio-assay with honeybee esterases and was expressed as an equivalent concentration of the model inhibitor dichlorvos. Estrogenic potency of the extracts was measured in the ER-CALUX reporter gene assay and was expressed as an equivalent concentration of estradiol. Multivariate principal component analysis (PCA) techniques proved to be valuable tools to analyze the numerous pesticide concentrations in relation to toxic potency, sampling location, and sampling season. Pesticide composition in rainwater depended much more on sampling season than on sampling location, but differences between SPRING and SUMMER were mainly attributed to local differences in agricultural practice. On average, the esterase inhibiting potency exceeded the maximum permissible concentration set for dichlorvos in The Netherlands, and was significantly higher in HORT than in BACK and BULB. Esterase inhibition correlated significantly with OP and CARB concentrations, as expected given the working mechanism of these insecticides. The estrogenic potency incidentally exceeded NOEC levels reported for

  6. The Chang'e 3 Mission Overview

    Science.gov (United States)

    Li, Chunlai; Liu, Jianjun; Ren, Xin; Zuo, Wei; Tan, Xu; Wen, Weibin; Li, Han; Mu, Lingli; Su, Yan; Zhang, Hongbo; Yan, Jun; Ouyang, Ziyuan

    2015-07-01

    The Chang'e 3 (CE-3) mission was implemented as the first lander/rover mission of the Chinese Lunar Exploration Program (CLEP). After its successful launch at 01:30 local time on December 2, 2013, CE-3 was inserted into an eccentric polar lunar orbit on December 6, and landed to the east of a 430 m crater in northwestern Mare Imbrium (19.51°W, 44.12°N) at 21:11 on December 14, 2013. The Yutu rover separated from the lander at 04:35, December 15, and traversed for a total of 0.114 km. Acquisition of science data began during the descent of the lander and will continue for 12 months during the nominal mission. The CE-3 lander and rover each carry four science instruments. Instruments on the lander are: Landing Camera (LCAM), Terrain Camera (TCAM), Extreme Ultraviolet Camera (EUVC), and Moon-based Ultraviolet Telescope (MUVT). The four instruments on the rover are: Panoramic Camera (PCAM), VIS-NIR Imaging Spectrometer (VNIS), Active Particle induced X-ray Spectrometer (APXS), and Lunar Penetrating Radar (LPR). The science objectives of the CE-3 mission include: (1) investigation of the morphological features and geological structures of and near the landing area; (2) integrated in-situ analysis of mineral and chemical composition of and near the landing area; and (3) exploration of the terrestrial-lunar space environment and lunar-based astronomical observations. This paper describes the CE-3 objectives and measurements that address the science objectives outlined by the Comprehensive Demonstration Report of Phase II of CLEP. The CE-3 team has archived the initial science data, and we describe data accessibility by the science community.

  7. A bacterial cocaine esterase protects against cocaine-induced epileptogenic activity and lethality.

    Science.gov (United States)

    Jutkiewicz, Emily M; Baladi, Michelle G; Cooper, Ziva D; Narasimhan, Diwahar; Sunahara, Roger K; Woods, James H

    2009-09-01

    Cocaine toxicity results in cardiovascular complications, seizures, and death and accounts for approximately 20% of drug-related emergency department visits every year. Presently, there are no treatments to eliminate the toxic effects of cocaine. The present study hypothesizes that a bacterial cocaine esterase with high catalytic efficiency would provide rapid and robust protection from cocaine-induced convulsions, epileptogenic activity, and lethality. Cocaine-induced paroxysmal activity and convulsions were evaluated in rats surgically implanted with radiotelemetry devices (N=6 per treatment group). Cocaine esterase was administered 1 minute after a lethal dose of cocaine or after cocaine-induced convulsions to determine the ability of the enzyme to prevent or reverse, respectively, the effects of cocaine. The cocaine esterase prevented all cocaine-induced electroencephalographic changes and lethality. This effect was specific for cocaine because the esterase did not prevent convulsions and death induced by a cocaine analog, (-)-2beta-carbomethoxy-3beta-phenyltropane. The esterase prevented lethality even after cocaine-induced convulsions occurred. In contrast, the short-acting benzodiazepine, midazolam, prevented cocaine-induced convulsions but not the lethal effects of cocaine. The data showed that cocaine esterase successfully degraded circulating cocaine to prevent lethality and that cocaine-induced convulsions alone are not responsible for the lethal effects of cocaine in this model. Therefore, further investigation into the use of cocaine esterase for treating cocaine overdose and its toxic effects is warranted.

  8. Functional classification of esterases from leaves of Aspidosperma polyneuron M. Arg. (Apocynaceae

    Directory of Open Access Journals (Sweden)

    Carvalho Vanda Marilza de

    2003-01-01

    Full Text Available Polyacrylamide gel electrophoresis system (PAGE and inhibition tests for biochemical characterization of alpha- and beta-esterases were used to obtain a functional classification of esterases fromAspidosperma polyneuron. The characterization of alpha- and beta-esterases from young leaves of A. polyneuron by the PAGE system showed fourteen esterase isozymes. The differential staining pattern showed that Est-2 isozyme hydrolyzes beta-naphthyl acetate; Est-6, Est-7 and Est-8 isozymes hydrolyze alpha-naphthyl acetate, and Est-1, Est-3, Est-4, Est-5, Est-9, Est-10, Est-11, Est-12, Est-13, and Est-14 isozymes hydrolyze both alpha- and b-naphthyl acetate. Inhibition pattern of a- and beta-esterases showed that Folidol is a more potent inhibitor that Malathion, while Thiamethoxan (an insecticide with organophosphorus-like action acts as an Est-4 and Est-6 inhibitor and induces the appearance of Est-5 and Est-7 isozymes as more intensely stained bands. Inhibition tests showed that OPC insecticides inhibit or activate plant esterases. Thus, plant esterases may be used as bioindicators to detect the presence and toxicity of residues of topically applied insecticides in agriculture and may be valuable for monitoring pollutants in the environment.

  9. A Novel Cold Active Esterase from a Deep Sea Sponge Stelletta normani Metagenomic Library

    Directory of Open Access Journals (Sweden)

    Erik Borchert

    2017-09-01

    Full Text Available Esterases catalyze the hydrolysis of ester bonds in fatty acid esters with short-chain acyl groups. Due to the widespread applications of lipolytic enzymes in various industrial applications, there continues to be an interest in novel esterases with unique properties. Marine ecosystems have long been acknowledged as a significant reservoir of microbial biodiversity and in particular of bacterial enzymes with desirable characteristics for industrial use, such as for example cold adaptation and activity in the alkaline pH range. We employed a functional metagenomic approach to exploit the enzymatic potential of one particular marine ecosystem, namely the microbiome of the deep sea sponge Stelletta normani. Screening of a metagenomics library from this sponge resulted in the identification of a number of lipolytic active clones. One of these encoded a highly, cold-active esterase 7N9, and the recombinant esterase was subsequently heterologously expressed in Escherichia coli. The esterase was classified as a type IV lipolytic enzyme, belonging to the GDSAG subfamily of hormone sensitive lipases. Furthermore, the recombinant 7N9 esterase was biochemically characterized and was found to be most active at alkaline pH (8.0 and displays salt tolerance over a wide range of concentrations. In silico docking studies confirmed the enzyme's activity toward short-chain fatty acids while also highlighting the specificity toward certain inhibitors. Furthermore, structural differences to a closely related mesophilic E40 esterase isolated from a marine sediment metagenomics library are discussed.

  10. A Lactobacillus plantarum Esterase Active on a Broad Range of Phenolic Esters

    Science.gov (United States)

    Esteban-Torres, María; Landete, José María; Reverón, Inés; Santamaría, Laura; de las Rivas, Blanca

    2015-01-01

    Lactobacillus plantarum is the lactic acid bacterial species most frequently found in the fermentation of food products of plant origin on which phenolic compounds are abundant. L. plantarum strains showed great flexibility in their ability to adapt to different environments and growth substrates. Of 28 L. plantarum strains analyzed, only cultures from 7 strains were able to hydrolyze hydroxycinnamic esters, such as methyl ferulate or methyl caffeate. As revealed by PCR, only these seven strains possessed the est_1092 gene. When the est_1092 gene was introduced into L. plantarum WCFS1 or L. lactis MG1363, their cultures acquired the ability to degrade hydroxycinnamic esters. These results support the suggestion that Est_1092 is the enzyme responsible for the degradation of hydroxycinnamic esters on the L. plantarum strains analyzed. The Est_1092 protein was recombinantly produced and biochemically characterized. Surprisingly, Est_1092 was able to hydrolyze not only hydroxycinnamic esters, since all the phenolic esters assayed were hydrolyzed. Quantitative PCR experiments revealed that the expression of est_1092 was induced in the presence of methyl ferulate, an hydroxycinnamic ester, but was inhibited on methyl gallate, an hydroxybenzoic ester. As Est_1092 is an enzyme active on a broad range of phenolic esters, simultaneously possessing feruloyl esterase and tannase activities, its presence on some L. plantarum strains provides them with additional advantages to survive and grow on plant environments. PMID:25746986

  11. A Lactobacillus plantarum esterase active on a broad range of phenolic esters.

    Science.gov (United States)

    Esteban-Torres, María; Landete, José María; Reverón, Inés; Santamaría, Laura; de las Rivas, Blanca; Muñoz, Rosario

    2015-05-01

    Lactobacillus plantarum is the lactic acid bacterial species most frequently found in the fermentation of food products of plant origin on which phenolic compounds are abundant. L. plantarum strains showed great flexibility in their ability to adapt to different environments and growth substrates. Of 28 L. plantarum strains analyzed, only cultures from 7 strains were able to hydrolyze hydroxycinnamic esters, such as methyl ferulate or methyl caffeate. As revealed by PCR, only these seven strains possessed the est_1092 gene. When the est_1092 gene was introduced into L. plantarum WCFS1 or L. lactis MG1363, their cultures acquired the ability to degrade hydroxycinnamic esters. These results support the suggestion that Est_1092 is the enzyme responsible for the degradation of hydroxycinnamic esters on the L. plantarum strains analyzed. The Est_1092 protein was recombinantly produced and biochemically characterized. Surprisingly, Est_1092 was able to hydrolyze not only hydroxycinnamic esters, since all the phenolic esters assayed were hydrolyzed. Quantitative PCR experiments revealed that the expression of est_1092 was induced in the presence of methyl ferulate, an hydroxycinnamic ester, but was inhibited on methyl gallate, an hydroxybenzoic ester. As Est_1092 is an enzyme active on a broad range of phenolic esters, simultaneously possessing feruloyl esterase and tannase activities, its presence on some L. plantarum strains provides them with additional advantages to survive and grow on plant environments. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  12. Studies on esterase isozymes and mycelium growth speed of ganoderma lucidum carried by Shenzhou spaceship

    International Nuclear Information System (INIS)

    Qi Jianjun; Chen Xiangdong; Lan Jin

    2002-01-01

    The esterase isozymes and mycelium growth speed of four Ganoderma lucidum strains carried by Shenzhou spaceship were studied. The results showed that different effects occurred to esterase and mycelium growth speed. The SX, S3 esterase band had changed compared with their control CX, C3, respectively, but there were no differences between SH and CH, S4 and C4. The growth speed of S4 strain was faster than its control C4, SX strain lower than its control CX, and there were no difference between SH and CH, S3 and C3

  13. G2E3 is a nucleo-cytoplasmic shuttling protein with DNA damage responsive localization

    International Nuclear Information System (INIS)

    Brooks, William S.; Banerjee, Sami; Crawford, David F.

    2007-01-01

    G2E3 was originally described as a G2/M-specific gene with DNA damage responsive expression. The presence of a conserved HECT domain within the carboxy-terminus of the protein indicated that it likely functions as a ubiquitin ligase or E3. Although HECT domains are known to function in this capacity for many proteins, we demonstrate that a portion of the HECT domain from G2E3 plays an important role in the dynamic subcellular localization of the protein. We have shown that G2E3 is a nucleo-cytoplasmic shuttling protein with nuclear export mediated by a novel nuclear export domain that functions independently of CRM1. In full-length G2E3, a separate region of the HECT domain suppresses the function of the NES. Additionally, G2E3 contains a nucleolar localization signal (NoLS) in its amino terminus. Localization of G2E3 to the nucleolus is a dynamic process, and the protein delocalizes from the nucleolus rapidly after DNA damage. Cell cycle phase-specific expression and highly regulated subcellular localization of G2E3 suggest a possible role in cell cycle regulation and the cellular response to DNA damage

  14. Isolation and Expression analysis of OsPME1, encoding for a putative Pectin Methyl Esterase from Oryza sativa (subsp. indica).

    Science.gov (United States)

    Kanneganti, Vydehi; Gupta, Aditya Kumar

    2009-04-01

    Pectin Methyl Esterases (PMEs) play an essential role during plant development by affecting the mechanical properties of the plant cell walls. Recent studies indicated that PMEs play important role in pollen tube development. In this study, we isolated a 1.3 kb cDNA clone from rice panicle cDNA library. It contained a 1038 bp of open reading frame (ORF) encoding for a putative pectin methyl esterase of 345 aminoacids with a 20 aminoacid signal peptide and was hence designated as OsPME1 (Oryza sativaPectin Methyl Esterase 1). It contained the structural arrangement GXYXE and GXXDFIF, found in the active groups of all PMEs. OsPME1 gene product shared varying identities, ranging from 52 % to 33 % with PMEs from other plant species belonging to Brassicaceae, Fabaceae, Amaranthaceae and Funariaceae. Southern blot analysis indicated that PME1 exists as a single copy in the rice genome. Expression pattern analysis revealed that OsPME1 is expressed only in pollen grains, during the later stages of their development and was also regulated by various abiotic stress treatments and phytohormones. Functional characterization of this pollen specific PME from rice would enable us to understand its role in pollen development.

  15. Plasma B-esterase activities in European raptors.

    Science.gov (United States)

    Roy, Claudie; Grolleau, Gérard; Chamoulaud, Serge; Rivière, Jean-Louis

    2005-01-01

    B-esterases are serine hydrolases composed of cholinesterases, including acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), and carboxylesterase (CbE). These esterases, found in blood plasma, are inhibited by organophosphorus (OP) and carbamate (CB) insecticides and can be used as nondestructive biomarkers of exposure to anticholinesterase insecticides. Furthermore, B-esterases are involved in detoxification of these insecticides. In order to establish the level of these enzymes and to have reference values for their normal activities, total plasma cholinesterase (ChE), AChE and BChE activities, and plasma CbE activity were determined in 729 European raptors representing 20 species, four families, and two orders. The diurnal families of the Falconiforme order were represented by Accipitridae and Falconidae and the nocturnal families of the Strigiforme order by Tytonidae and Strigidae. Intraspecies differences in cholinesterase activities according to sex and/or age were investigated in buzzards (Buteo buteo), sparrowhawks (Accipiter nisus), kestrels (Falco tinnunculus), barn owls (Tyto alba), and tawny owls (Strix aluco). Sex-related differences affecting ChE and AChE activities were observed in young kestrels (2-3-mo-old) and age-related differences in kestrels (ChE and AChE), sparrowhawks (AChE), and tawny owls (ChE, AChE, and BChE). The interspecies analysis yielded a negative correlation between ChE activity and body mass taking into account the relative contribution of AChE and BChE to ChE activity, with the exception of the honey buzzard (Pernis apivorus). The lowest ChE activities were found in the two largest species, Bonelli's eagle (Hieraaetus fasciatus) and Egyptian vulture (Neophron percnopterus) belonging to the Accipitridae family. The highest ChE activities were found in the relatively small species belonging to the Tytonidae and Strigidae families and in honey buzzard of the Accipitridae family. Species of the Accipitridae, Tytonidae, and

  16. Esterases no exame da estrutura populacional de Camu-camu (Myrciaria dubia (Kunth McVaugh-Myrtaceae Esterases for examining the population structure of Camu-camu (Myrciaria dubia (Kunth McVaugh-Myrtaceae

    Directory of Open Access Journals (Sweden)

    Aylton Saturnino Teixeira

    2004-01-01

    populacional de Uatumã, devem-se ao fato dessa amostra ter apresentado um loco polimórfico a mais (Est-D2, em relação as demais amostras investigadas.Two enzymatic systems (esterase and esterase-D, analysed by using the starch gel electrophoresis technique on plant young leaves cultivated in the higher land (terra firme, from seeds of camu-camu, Myrciaria dubia (Kunth McVaugh-Myrtaceae obtained from three natural population samples (Iquitos, Boa Vista and Uatumã, revealed 6 loci: Est-1, Est-2, Est-3, Est-4, Est-D1 and Est-D2. Two of the six gene loci presently examined (Est-3 and Est-D2 showed to be polymorphic, so they happen to be valuable for characterizing the species population structure. The polymorphism patterns shown in the Est-3 and Est-D2 loci of camu-camu, are typical of monomeric and dimeric enzymes, respectively. The locus Est-3 showed a huge genetic imbalance within and among the population samples examined, due to an excessive observed numbers of heterozygote plants compared to their expected numbers. The locus Est-D2 showed an exclusive polymorphism for the alleles Est-D2¹, Est-D2² and Est-D2³, and a good genetic balance in the Uatumã population sample. On account of that, among all gene loci investigated here, Est-D2 seems to be the most suitable locus for identifying and delimiting camu-camu probable stocks. Hence, this locus has to be present in the list of the isoenzymatic markers for future research on the species population genetics in the Amazon region. Data on allelic frequency distribution, genetic distance and genetic variation estimates on the 6 esterase loci investigated, were effective for demonstrating genetic differences among the surveyed species population samples. The highest values of mean heterozigozity (0.1353; proportion of polimorphic loci (0.33 and average number of alleles per locus (1.33 revealed in the Uatumã population sample, are due to the presence of an extra polymorphic locus (Est-D2 when compared with other surveyed

  17. Fitness differences due to allelic variation at Esterase-4 locus in ...

    Indian Academy of Sciences (India)

    KAVITA KRISHNAMOORTI

    2017-08-31

    Aug 31, 2017 ... Keywords. esterases; null allele; reproductive fitness; natural selection; Drosophila ananassae. .... cific substrate (1-naphthylacetate AR) and stain (fast blue. RR). On the ... transferred to fresh food vials and eggs were counted.

  18. Common and Distant Structural Characteristics of Feruloyl Esterase Families from Aspergillus oryzae

    DEFF Research Database (Denmark)

    Udatha, D. B. R. K. Gupta; Mapelli, Valeria; Panagiotou, Gianni

    2012-01-01

    Background: Feruloyl esterases (FAEs) are important biomass degrading accessory enzymes due to their capability of cleaving the ester links between hemicellulose and pectin to aromatic compounds of lignin, thus enhancing the accessibility of plant tissues to cellulolytic and hemicellulolytic...

  19. Multiple nucleophilic elbows leading to multiple active sites in a single module esterase from Sorangium cellulosum

    DEFF Research Database (Denmark)

    Udatha, D.B.R.K. Gupta; Madsen, Karina Marie; Panagiotou, Gianni

    2015-01-01

    The catalytic residues in carbohydrate esterase enzyme families constitute a highly conserved triad: serine, histidine and aspartic acid. This catalytic triad is generally located in a very sharp turn of the protein backbone structure, called the nucleophilic elbow and identified by the consensus...... sequence GXSXG. An esterase from Sorangium cellulosum Soce56 that contains five nucleophilic elbows was cloned and expressed in Escherichia coli and the function of each nucleophilic elbowed site was characterized. In order to elucidate the function of each nucleophilic elbow, site directed mutagenesis....... To our knowledge, this is the first report presenting the role of multiple nucleophilic elbows in the catalytic promiscuity of an esterase. Further structural analysis at protein unit level indicates the new evolutionary trajectories in emerging promiscuous esterases....

  20. Esterase-sensitive sulfur dioxide prodrugs inspired by modified Julia olefination.

    Science.gov (United States)

    Wang, Wenyi; Wang, Binghe

    2017-09-12

    Sulfur dioxide (SO 2 ) is an endogenously produced gaseous molecule, and is emerging as a potential gasotransmitter. Herein, we describe the first series of esterase-sensitive prodrugs inspired by modified Julia olefination as SO 2 donors.

  1. Esterase-D and chromosome patterns in Central Amazon piranha (Serrasalmus rhombeus Linnaeus, 1766 from Lake Catalão

    Directory of Open Access Journals (Sweden)

    Aylton Saturnino Teixeira

    2006-01-01

    Full Text Available This study presents additional genetic data on piranha (Serrasalmus rhombeus Linnaeus, 1766 complex previously diagnosed due to the presence of distinct cytotypes 2n = 58 and 2n = 60. Three esterase-D enzyme loci (Est-D1, Est-D2 and Est-D3 were examined and complemented with chromosomal data from 66 piranha specimens collected from Lake Catalão. For all specimens the Est-D1 and Est-D2 loci were monomorphic. In contrast, the Est-D3 locus was polymorphic with genotypes and alleles being differentially distributed in the previously described cytotypes and served as the basis for detecting a new cytotype (2n = 60 B. In cytotype 2n = 58 the Est-D3 locus was also polymorphic and presented Mendelian allelic segregation with four genotypes (Est-D3(11, Est-D3(12, Est-D3(22 and Est-D3(33 out of six theoretically possible genotypes, presumably encoded by alleles Est-D3¹ (frequency = 0.237, EsT-D3² (0.710 and Est-D3³ (0.053. A Chi-squared (chi2 test for Hardy-Weinberg equilibrium was applied to the Est-D3 locus and revealed a genetic unbalance in cytotype 2n = 58, indicating the probable existence in the surveyed area of different stocks for that karyotypic structure. A silent null allele (Est-D3(0 with a high frequency (0.959 occurred exclusively in the 2n = 60 cytotype. On the other hand, the new cytotype 2n = 60 B described here for the first time was monomorphic for the presumably fixed Est-D3³ allele. The data as a whole should contribute to the better understanding the rhombeus complex taxonomic status definition in the Central Amazon.

  2. Serine esterase and hemolytic activity in human cloned cytotoxic T lymphocytes

    OpenAIRE

    1988-01-01

    Target cell lysis by most murine cytotoxic T lymphocytes appears to be mediated by a complement (C9)-like protein called perforin, contained in high-density cytoplasmic granules. These granules also contain high levels of serine esterase activity, which may also play a role in cytolysis. Analysis of 17 cloned human cytotoxic T lymphocytes revealed the presence of serine esterase that is very similar to its murine counterpart in substrate and inhibitor specificities, pH optimum, and molecular ...

  3. The search of the target of promotion: Phenylbenzoate esterase activities in hen peripheral nerve

    International Nuclear Information System (INIS)

    Moretto, A.; Nicolli, A.; Lotti, M.

    2007-01-01

    Certain esterase inhibitors, such as carbamates, phosphinates and sulfonyl halides, do not cause neuropathy as some organophosphates, but they may exacerbate chemical or traumatic insults to axons. This phenomenon is called promotion of axonopathies. Given the biochemical and toxicological characteristics of these compounds, the hypothesis was made that the target of promotion is a phenyl valerate (PV) esterase similar to neuropathy target esterase (NTE), the target of organophosphate induced delayed polyneuropathy. However, attempts to identify a PV esterase in hen peripheral nerve have been, so far, unsuccessful. We tested several esters, other than PV, as substrates of esterases from crude homogenate of the hen peripheral nerve. The ideal substrate should be poorly hydrolysed by NTE but extensively by enzyme(s) that are insensitive to non-promoters, such as mipafox, and sensitive to promoters, such as phenyl methane sulfonyl fluoride (PMSF). When phenyl benzoate (PB) was used as substrate, about 65% of total activity was resistant to the non-promoter mipafox (up to 0.5 mM, 20 min, pH 8.0), that inhibits NTE and other esterases. More than 90% of this resistant activity was sensitive to the classical promoter PMSF (1 mM, 20 min, pH 8.0) with an IC 50 of about 0.08 mM (20 min, pH 8.0). On the contrary, the non-promoter p-toluene sulfonyl fluoride caused only about 10% inhibition at 0.5 mM. Several esterase inhibitors including, paraoxon, phenyl benzyl carbamate, di-n-butyl dichlorovinyl phosphate and di-isopropyl fluorophosphate, were tested both in vitro and in vivo for inhibition of this PB activity. Mipafox-resistant PMSF-sensitive PB esterase activity(ies) was inhibited by promoters but not by non promoters and neuropathic compounds

  4. Expression and characterization of a new esterase with GCSAG motif from a permafrost metagenomic library.

    Science.gov (United States)

    Petrovskaya, Lada E; Novototskaya-Vlasova, Ksenia A; Spirina, Elena V; Durdenko, Ekaterina V; Lomakina, Galina Yu; Zavialova, Maria G; Nikolaev, Evgeny N; Rivkina, Elizaveta M

    2016-05-01

    As a result of construction and screening of a metagenomic library prepared from a permafrost-derived microcosm, we have isolated a novel gene coding for a putative lipolytic enzyme that belongs to the hormone-sensitive lipase family. It encodes a polypeptide of 343 amino acid residues whose amino acid sequence displays maximum likelihood with uncharacterized proteins from Sphingomonas species. A putative catalytic serine residue of PMGL2 resides in a new variant of a recently discovered GTSAG sequence in which a Thr residue is replaced by a Cys residue (GCSAG). The recombinant PMGL2 was produced in Escherichia coli cells and purified by Ni-affinity chromatography. The resulting protein preferably utilizes short-chain p-nitrophenyl esters (C4 and C8) and therefore is an esterase. It possesses maximum activity at 45°C in slightly alkaline conditions and has limited thermostability at higher temperatures. Activity of PMGL2 is stimulated in the presence of 0.25-1.5 M NaCl indicating the good salt tolerance of the new enzyme. Mass spectrometric analysis demonstrated that N-terminal methionine in PMGL2 is processed and cysteine residues do not form a disulfide bond. The results of the study demonstrate the significance of the permafrost environment as a unique genetic reservoir and its potential for metagenomic exploration. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. HIGHLY METHYL ESTERIFIED SEEDS is a pectin methyl esterase involved in embryo development.

    Science.gov (United States)

    Levesque-Tremblay, Gabriel; Müller, Kerstin; Mansfield, Shawn D; Haughn, George W

    2015-03-01

    Homogalacturonan pectin domains are synthesized in a highly methyl-esterified form that later can be differentially demethyl esterified by pectin methyl esterase (PME) to strengthen or loosen plant cell walls that contain pectin, including seed coat mucilage, a specialized secondary cell wall of seed coat epidermal cells. As a means to identify the active PMEs in seed coat mucilage, we identified seven PMEs expressed during seed coat development. One of these, HIGHLY METHYL ESTERIFIED SEEDS (HMS), is abundant during mucilage secretion, peaking at 7 d postanthesis in both the seed coat and the embryo. We have determined that this gene is required for normal levels of PME activity and homogalacturonan methyl esterification in the seed. The hms-1 mutant displays altered embryo morphology and mucilage extrusion, both of which are a consequence of defects in embryo development. A significant decrease in the size of cells in the embryo suggests that the changes in embryo morphology are a consequence of lack of cell expansion. Progeny from a cross between hms-1 and the previously characterized PME inhibitor5 overexpression line suggest that HMS acts independently from other cell wall-modifying enzymes in the embryo. We propose that HMS is required for cell wall loosening in the embryo to facilitate cell expansion during the accumulation of storage reserves and that its role in the seed coat is masked by redundancy. © 2015 American Society of Plant Biologists. All Rights Reserved.

  6. Esterase inhibition by synergists in the western flower thrips Frankliniella occidentalis.

    Science.gov (United States)

    López-Soler, Neus; Cervera, Amelia; Quinto, Vicente; Abellán, Jaime; Bielza, Pablo; Martínez-Pardo, Rafael; Garcerá, Maria Dolores

    2011-12-01

    Western flower thrips (WFT), Frankliniella occidentalis (Pergande), is among the most important crop pests in the south-eastern region of Spain. Its increasing resistance to insecticides constitutes a serious problem, and understanding the mechanisms involved is therefore of great interest. Use of synergists to inhibit the enzymes involved in insecticide detoxification is widely used to determine their responsibility for insecticide resistance. However, they do not always act as intended or expected, and caution must be exercised when interpreting synergist results. Laboratory-selected strains of WFT were used to analyse the effects of the synergists piperonyl butoxide (PBO), S,S,S-tributyl phosphorotrithioate (DEF) and methiocarb on total esterase activity. Significant differences were found, indicating esterase activity inhibition by DEF, a lower effect for methiocarb and a small inhibition of the activity by PBO. Esterase isoenzyme inhibition by these compounds showed a similar result; this assay revealed an extreme sensitivity of Triplet A (resistance-associated esterases) to DEF. In an in vivo assay carried out with these compounds at different incubation times, only DEF caused posterior in vitro esterase activity inhibition, with a maximum effect 1 h after treatment. In this work, only DEF shows true synergistic inhibition of WFT esterases. Copyright © 2011 Society of Chemical Industry.

  7. Depletion of juvenile hormone esterase extends larval growth in Bombyx mori.

    Science.gov (United States)

    Zhang, Zhongjie; Liu, Xiaojing; Shiotsuki, Takahiro; Wang, Zhisheng; Xu, Xia; Huang, Yongping; Li, Muwang; Li, Kai; Tan, Anjiang

    2017-02-01

    Two major hormones, juvenile hormone (JH) and 20-hydroxyecdysone (20E), regulate insect growth and development according to their precisely coordinated titres, which are controlled by both biosynthesis and degradation pathways. Juvenile hormone esterase (JHE) is the primary JH-specific degradation enzyme that plays a key role in regulating JH titers, along with JH epoxide hydrolase (JHEH) and JH diol kinase (JHDK). In the current study, a loss-of-function analysis of JHE in the silkworm, Bombyx mori, was performed by targeted gene disruption using the transgenic CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/RNA-guided Cas9 nucleases) system. Depletion of B. mori JHE (BmJHE) resulted in the extension of larval stages, especially the penultimate and ultimate larval stages, without deleterious effects to silkworm physiology. The expression of JHEH and JHDK was upregulated in mutant animals, indicating the existence of complementary routes in the JH metabolism pathway in which inactivation of one enzyme will activate other enzymes. RNA-Seq analysis of mutant animals revealed that genes involved in protein processing in the endoplasmic reticulum and in amino acid metabolism were affected by BmJHE depletion. Depletion of JHE and subsequent delayed JH metabolism activated genes in the TOR pathway, which are ultimately responsible for extending larval growth. The transgenic Cas9 system used in the current study provides a promising approach for analysing the actions of JH, especially in nondrosophilid insects. Furthermore, prolonging larval stages produced larger larvae and cocoons, which is greatly beneficial to silk production. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Characterization of a cold-active esterase from Serratia sp. and improvement of thermostability by directed evolution.

    Science.gov (United States)

    Jiang, Huang; Zhang, Shaowei; Gao, Haofeng; Hu, Nan

    2016-01-22

    In recent years, cold-active esterases have received increased attention due to their attractive properties for some industrial applications such as high catalytic activity at low temperatures. An esterase-encoding gene (estS, 909 bp) from Serratia sp. was identified, cloned and expressed in Escherichia coli DE3 (BL21). The estS encoded a protein (EstS) of 302 amino acids with a predicted molecular weight of 32.5 kDa. It showed the highest activity at 10 °C and pH 8.5. EstS was cold active and retained ~92 % of its original activity at 0 °C. Thermal inactivation analysis showed that the T1/2 value of EstS was 50 min at 50 °C (residual activity 41.23 %) after 1 h incubation. EstS is also quite stable in high salt conditions and displayed better catalytic activity in the presence of 4 M NaCl. To improve the thermo-stability of EstS, variants of estS gene were created by error-prone PCR. A mutant 1-D5 (A43V, R116W, D147N) that showed higher thermo-stability than its wild type predecessor was selected. 1-D5 showed enhanced T1/2 of 70 min at 50 °C and retained 63.29 % of activity after incubation at 50 °C for 60 min, which were about 22 % higher than the wild type (WT). CD spectrum showed that the secondary structure of WT and 1-D5 are more or less similar, but an increase in β-sheets was recorded, which enhanced the thermostability of mutant protein. EstS was a novel cold-active and salt-tolerant esterase and half-life of mutant 1-D5 was enhanced by 1.4 times compared with WT. The features of EstS are interesting and can be exploited for commercial applications. The results have also provided useful information about the structure and function of Est protein.

  9. Switching catalysis from hydrolysis to perhydrolysis in P. fluorescens esterase

    Science.gov (United States)

    Yin, De Lu (Tyler); Bernhardt, Peter; Morley, Krista L.; Jiang, Yun; Cheeseman, Jeremy D.; Purpero, Vincent; Schrag, Joseph D.; Kazlauskas, Romas J.

    2010-01-01

    Many serine hydrolases catalyze perhydrolysis – the reversible formation of per-acids from carboxylic acids and hydrogen peroxide. Recently we showed that a single amino acid substitution in the alcohol binding pocket - L29P - in Pseudomonas fluorescens (SIK WI) aryl esterase (PFE) increased the specificity constant of PFE for peracetic acid formation >100-fold [Bernhardt et al. Angew. Chem. Intl. Ed. 2005, 44, 2742]. In this paper, we extend this work to address the three following questions. First, what is the molecular basis of the increase in perhydrolysis activity? We previously proposed that the L29P substitution creates a hydrogen bond between the enzyme and hydrogen peroxide in the transition state. Here we report two x-ray structures of L29P PFE that support this proposal. Both structures show a main chain carbonyl oxygen closer to the active-site serine as expected. One structure further shows acetate in the active site in an orientation consistent with reaction by an acyl-enzyme mechanism. We also detected an acyl-enzyme intermediate in the hydrolysis of ε-caprolactone by mass spectrometry. Second, can we further increase perhydrolysis activity? We discovered that the reverse reaction – hydrolysis of peracetic acid to acetic acid and hydrogen peroxide – occurs at nearly the diffusion limited rate. Since the reverse reaction cannot increase further, neither can the forward reaction. Consistent with this prediction, two variants with additional amino acid substitutions showed two fold higher kcat, but Km also increased so the specificity constant, kcat/Km, remained similar. Third, how does the L29P substitution change the esterase activity? Ester hydrolysis decreased for most esters (75-fold for ethyl acetate), but not for methyl esters. In contrast, L29P PFE catalyzed hydrolysis of ε-caprolactone five times more efficiently than wild-type PFE. Molecular modeling suggests that moving the carbonyl group closer to the active site blocks access for

  10. Switching Catalysis from Hydrolysis to Perhydrolysis in Pseudomonas fluorescens Esterase

    Energy Technology Data Exchange (ETDEWEB)

    Yin, D.; Bernhardt, P; Morley, K; Jiang, Y; Cheeseman, J; Purpero, V; Schrag, J; Kazlauskas, R

    2010-01-01

    Many serine hydrolases catalyze perhydrolysis, the reversible formation of peracids from carboxylic acids and hydrogen peroxide. Recently, we showed that a single amino acid substitution in the alcohol binding pocket, L29P, in Pseudomonas fluorescens (SIK WI) aryl esterase (PFE) increased the specificity constant of PFE for peracetic acid formation >100-fold [Bernhardt et al. (2005) Angew. Chem., Int. Ed. 44, 2742]. In this paper, we extend this work to address the three following questions. First, what is the molecular basis of the increase in perhydrolysis activity? We previously proposed that the L29P substitution creates a hydrogen bond between the enzyme and hydrogen peroxide in the transition state. Here we report two X-ray structures of L29P PFE that support this proposal. Both structures show a main chain carbonyl oxygen closer to the active site serine as expected. One structure further shows acetate in the active site in an orientation consistent with reaction by an acyl-enzyme mechanism. We also detected an acyl-enzyme intermediate in the hydrolysis of {var_epsilon}-caprolactone by mass spectrometry. Second, can we further increase perhydrolysis activity? We discovered that the reverse reaction, hydrolysis of peracetic acid to acetic acid and hydrogen peroxide, occurs at nearly the diffusion limited rate. Since the reverse reaction cannot increase further, neither can the forward reaction. Consistent with this prediction, two variants with additional amino acid substitutions showed 2-fold higher k{sub cat}, but K{sub m} also increased so the specificity constant, k{sub cat}/K{sub m}, remained similar. Third, how does the L29P substitution change the esterase activity? Ester hydrolysis decreased for most esters (75-fold for ethyl acetate) but not for methyl esters. In contrast, L29P PFE catalyzed hydrolysis of {var_epsilon}-caprolactone five times more efficiently than wild-type PFE. Molecular modeling suggests that moving the carbonyl group closer to the

  11. Overexpression of herbaceous peony miR156e-3p improves anthocyanin accumulation in transgenic Arabidopsis thaliana lateral branches.

    Science.gov (United States)

    Zhao, Daqiu; Xia, Xing; Wei, Mengran; Sun, Jing; Meng, Jiasong; Tao, Jun

    2017-12-01

    microRNAs (miRNAs) play critical regulatory roles in plant growth and development. In the present study, the function of herbaceous peony ( Paeonia lactiflora Pall.) miR156e-3p in the regulation of color formation has been investigated. Firstly, P. lactiflora miR156e-3p precursor sequence (pre-miR156e-3p) was isolated. Subsequently, the overexpression vector of pre-miR156e-3p was constructed and transformed into Arabidopsis thaliana . Moreover, the medium screening, GUS staining, polymerase chain reaction (PCR) of the GUS region and real-time quantitative PCR (qRT-PCR) of miR156e-3p all confirmed that the purpose gene had been successfully transferred into Arabidopsis plants and expressed, which resulted in apparent purple lateral branches. And this change in color was caused by the improved anthocyanin accumulation. In addition, expression analysis had shown that the level of miR156e-3p transcript was increased, while transcription level of target gene squamosa promoter binding protein-like gene ( SPL1 ), encoding SPL transcription factor that negatively regulated anthocyanin accumulation, was repressed in miR156e-3p-overexpressing transgenic plants, and its downstream gene dihydroflavonol 4-reductase gene ( DFR ) that was directly involved in anthocyanin biosynthesis was strongly expressed, which resulted in anthocyanin accumulation of Arabidopsis lateral branches. These findings would improve the understanding of miRNAs regulation of color formation in P. lactiflora .

  12. Structural analysis of thermostabilizing mutations of cocaine esterase

    Energy Technology Data Exchange (ETDEWEB)

    Narasimhan, Diwahar; Nance, Mark R.; Gao, Daquan; Ko, Mei-Chuan; Macdonald, Joanne; Tamburi, Patricia; Yoon, Dan; Landry, Donald M.; Woods, James H.; Zhan, Chang-Guo; Tesmer, John J.G.; Sunahara, Roger K. (Michigan); (Columbia); (Kentucky)

    2010-09-03

    Cocaine is considered to be the most addictive of all substances of abuse and mediates its effects by inhibiting monoamine transporters, primarily the dopamine transporters. There are currently no small molecules that can be used to combat its toxic and addictive properties, in part because of the difficulty of developing compounds that inhibit cocaine binding without having intrinsic effects on dopamine transport. Most of the effective cocaine inhibitors also display addictive properties. We have recently reported the use of cocaine esterase (CocE) to accelerate the removal of systemic cocaine and to prevent cocaine-induced lethality. However, wild-type CocE is relatively unstable at physiological temperatures ({tau}{sub 1/2} {approx} 13 min at 37 C), presenting challenges for its development as a viable therapeutic agent. We applied computational approaches to predict mutations to stabilize CocE and showed that several of these have increased stability both in vitro and in vivo, with the most efficacious mutant (T172R/G173Q) extending half-life up to 370 min. Here we present novel X-ray crystallographic data on these mutants that provide a plausible model for the observed enhanced stability. We also more extensively characterize the previously reported variants and report on a new stabilizing mutant, L169K. The improved stability of these engineered CocE enzymes will have a profound influence on the use of this protein to combat cocaine-induced toxicity and addiction in humans.

  13. Acetylcholine esterase activity in mild cognitive impairment and Alzheimer's disease

    International Nuclear Information System (INIS)

    Herholz, Karl

    2008-01-01

    Impairment of cholinergic neurotransmission is a well-established fact in Alzheimer's disease (AD), but there is controversy about its relevance at the early stages of the disease and in mild cognitive impairment (MCI). In vivo positron emission tomography imaging of cortical acetylcholine esterase (AChE) activity as a marker of cholinergic innervation that is expressed by cholinergic axons and cholinoceptive neurons has demonstrated a reduction of this enzyme activity in manifest AD. The technique is also useful to measure the inhibition of cerebral AChE induced by cholinesterase inhibitors for treatment of dementia symptoms. A reduction of cortical AchE activity was found consistently in all studies of AD and in few cases of MCI who later concerted to AD. The in vivo findings in MCI and very mild AD are still preliminary, and studies seem to suggest that cholinergic innervation and AChE as the main degrading enzyme are both reduced, which might result in partial compensation of their effect. (orig.)

  14. Gender differences in the activities of aspirin-esterases in rat tissues

    Directory of Open Access Journals (Sweden)

    Benedito M.A.C.

    1998-01-01

    Full Text Available The activities of aspirin (acetylsalicylic acid-esterases were measured in several tissues (liver, kidney, adrenal glands, brain and serum from adult male and female Wistar rats. In males, both aspirin-esterase I (assayed at pH 5.5 and II (assayed at pH 7.4 activities were higher in liver homogenates when compared to females (aspirin-esterase I: males 48.9 ± 4.8 (N = 8 and females 29.3 ± 4.2 (N = 8 nmol of salicylic acid formed min-1 mg protein-1; aspirin-esterase II: males 41.4 ± 4.1 (N = 8 and females 26.1 ± 4.5 (N = 8 nmol of salicylic acid formed min-1 mg protein-1, P<0.001. In serum, enzyme activity was higher in females than in males (aspirin-esterase I: males 0.85 ± 0.06 (N = 6 and females 1.18 ± 0.11 (N = 6 nmol of salicylic acid formed min-1 mg protein-1; aspirin-esterase II: males 1.03 ± 0.13 (N = 6 and females 1.34 ± 0.11 (N = 6 nmol of salicylic acid formed min-1 mg protein-1, P<0.001. In the other tissues assayed, no statistically significant difference between males and females was found. There were no statistically significant differences when the enzymes were assayed in different phases of the estrous cycle in liver and serum. These results show that the differences in aspirin-esterase activity observed between males and females are not due to the estrous cycle. The gender difference obtained in our study may indicate an involvement of gonadal hormones in the control of the hydrolysis of aspirin. This possibility is currently under investigation.

  15. Some nonunitary, indecomposable representations of the Euclidean algebra e(3)

    International Nuclear Information System (INIS)

    Douglas, Andrew; De Guise, Hubert

    2010-01-01

    The Euclidean group E(3) is the noncompact, semidirect product group E(3)≅R 3 x SO(3). It is the Lie group of orientation-preserving isometries of three-dimensional Euclidean space. The Euclidean algebra e(3) is the complexification of the Lie algebra of E(3). We construct three distinct families of finite-dimensional, nonunitary representations of e(3) and show that each representation is indecomposable. The representations of the first family are explicitly realized as subspaces of the polynomial ring F[X,Y,Z] with the action of e(3) given by differential operators. The other families are constructed via duals and tensor products of the representations within the first family. We describe subrepresentations, quotients and duals of these indecomposable representations.

  16. Solid-state fermentation as a potential technique for esterase/lipase production by halophilic archaea.

    Science.gov (United States)

    Martin del Campo, Martha; Camacho, Rosa M; Mateos-Díaz, Juan C; Müller-Santos, Marcelo; Córdova, Jesus; Rodríguez, Jorge A

    2015-11-01

    Halophilic archaea are extremophiles, adapted to high-salt environments, showing a big biotechnological potential as enzyme, lipids and pigments producers. Four inert supports (perlite, vermiculite, polyurethane foam and glass fiber) were employed for solid-state fermentation (SSF) of the halophilic archaeon Natronococcus sp. TC6 to investigate biomass and esterase production. A very low esterase activity and high water activity were observed when perlite, vermiculite and polyurethane were used as supports. When glass fiber was employed, an important moisture loss was observed (8.6%). Moreover, moisture retention was improved by mixing polyurethane and glass fiber, resulting in maximal biomass and esterase production. Three halophilic archaea: Natronococcus sp. TC6, Halobacterium sp. NRC-1 and Haloarcula marismortui were cultured by submerged fermentation (SmF) and by SSF; an improvement of 1.3- to 6.2-fold was observed in the biomass and esterase production when SSF was used. Growth was not homogeneous in the mixture, but was predominant in the glass fiber thus was probably because the glass fiber provides a holder to the cells, while the polyurethane acts as an impregnation medium reservoir. To the best of our knowledge, this work is the first report on haloarchaea cultivation by SSF aiming biomass and esterase/lipase activity production.

  17. Esterase Active in Polar Organic Solvents from the Yeast Pseudozyma sp. NII 08165

    Directory of Open Access Journals (Sweden)

    Deepthy Alex

    2014-01-01

    Full Text Available Esterases/lipases active in water miscible solvents are highly desired in biocatalysis where substrate solubility is limited and also when the solvent is desired as an acyl acceptor in transesterification reactions, as with the case of biodiesel production. We have isolated an esterase from the glycolipid producing yeast-Pseudozyma sp. NII 08165 which in its crude form was alkali active, thermo stable, halo tolerant and also capable of acting in presence of high methanol concentration. The crude enzyme which maintained 90% of its original activity after being treated at 70°C was purified and the properties were characterized. The partially purified esterase preparation had temperature and pH optima of 60°C and 8.0 respectively. The enzyme retained almost complete activity in presence of 25% methanol and 80% activity in the same strength of ethanol. Conditions of enzyme production were optimized, which lead to 9 fold increase in the esterase yield. One of the isoforms of the enzyme LIP1 was purified to homogeneity and characterized. Purified LIP1 had a Km and Vmax of 0.01 and 1.12, respectively. The purified esterase lost its thermo and halo tolerance but interestingly, retained 97% activity in methanol.

  18. Establishment of a Wheat Cell-Free Synthesized Protein Array Containing 250 Human and Mouse E3 Ubiquitin Ligases to Identify Novel Interaction between E3 Ligases and Substrate Proteins.

    Directory of Open Access Journals (Sweden)

    Hirotaka Takahashi

    Full Text Available Ubiquitination is a key post-translational modification in the regulation of numerous biological processes in eukaryotes. The primary roles of ubiquitination are thought to be the triggering of protein degradation and the regulation of signal transduction. During protein ubiquitination, substrate specificity is mainly determined by E3 ubiquitin ligase (E3. Although more than 600 genes in the human genome encode E3, the E3s of many target proteins remain unidentified owing to E3 diversity and the instability of ubiquitinated proteins in cell. We demonstrate herein a novel biochemical analysis for the identification of E3s targeting specific proteins. Using wheat cell-free protein synthesis system, a protein array containing 227 human and 23 mouse recombinant E3s was synthesized. To establish the high-throughput binding assay using AlphaScreen technology, we selected MDM2 and p53 as the model combination of E3 and its target protein. The AlphaScreen assay specifically detected the binding of p53 and MDM2 in a crude translation mixture. Then, a comprehensive binding assay using the E3 protein array was performed. Eleven of the E3s showed high binding activity, including four previously reported E3s (e.g., MDM2, MDM4, and WWP1 targeting p53. This result demonstrated the reliability of the assay. Another interactors, RNF6 and DZIP3-which there have been no report to bind p53-were found to ubiquitinate p53 in vitro. Further analysis showed that RNF6 decreased the amount of p53 in H1299 cells in E3 activity-dependent manner. These results suggest the possibility that the RNF6 ubiquitinates and degrades p53 in cells. The novel in vitro screening system established herein is a powerful tool for finding novel E3s of a target protein.

  19. Cellular function of neuropathy target esterase in lysophosphatidylcholine action

    International Nuclear Information System (INIS)

    Vose, Sarah C.; Fujioka, Kazutoshi; Gulevich, Alex G.; Lin, Amy Y.; Holland, Nina T.; Casida, John E.

    2008-01-01

    Neuropathy target esterase (NTE) plays critical roles in embryonic development and maintenance of peripheral axons. It is a secondary target of some organophosphorus toxicants including analogs of insecticides and chemical warfare agents. Although the mechanistic role of NTE in vivo is poorly defined, it is known to hydrolyze lysophosphatidylcholine (LPC) in vitro and may protect cell membranes from cytotoxic accumulation of LPC. To determine the cellular function of NTE, Neuro-2a and COS-7 cells were transfected with a full-length human NTE-containing plasmid yielding recombinant NTE (rNTE). We find the same inhibitor sensitivity and specificity profiles for rNTE assayed with LPC or phenyl valerate (a standard NTE substrate) and that this correlation extends to the LPC hydrolases of human brain, lymphocytes and erythrocytes. All of these LPC hydrolases are therefore very similar to each other in respect to a conserved inhibitor binding site conformation. NTE is expressed in brain and lymphocytes and contributes to LPC hydrolase activities in these tissues. The enzyme or enzymes responsible for erythrocyte LPC hydrolase activity remain to be identified. We also show that rNTE protects Neuro-2a and COS-7 cells from exogenous LPC cytotoxicity. Expression of rNTE in Neuro-2a cells alters their phospholipid balance (analyzed by liquid chromatography-mass spectrometry with single ion monitoring) by lowering LPC-16:0 and LPC-18:0 and elevating glycerophosphocholine without a change in phosphatidylcholine-16:0/18:1 or 16:0/18:2. NTE therefore serves an important function in LPC homeostasis and action

  20. Characterization and mode of action of two acetyl xylan esterases from Chrysosporium lucknowense C1 active towards acetylated xylans

    NARCIS (Netherlands)

    Pouvreau, L.A.M.; Jonathan, M.C.; Kabel, M.A.; Hinz, S.W.A.; Gruppen, H.; Schols, H.A.

    2011-01-01

    Two novel acetyl xylan esterases, Axe2 and Axe3, from Chrysosporium lucknowense (C1), belonging to the carbohydrate esterase families 5 and 1, respectively, were purified and biochemically characterized. Axe2 and Axe3 are able to hydrolyze acetyl groups both from simple acetylated

  1. Enzymatic degradation of lignin‐carbohydrate complexes (LCCs): Model studies using a fungal glucuronoyl esterase from Cerrena unicolor

    DEFF Research Database (Denmark)

    d'Errico, Clotilde; Jørgensen, Jonas O.; Krogh, Kristian B. R. M.

    2015-01-01

    Lignin‐carbohydrate complexes (LCCs) are believed to influence the recalcitrance of lignocellulosic plant material preventing optimal utilization of biomass in e.g. forestry, feed and biofuel applications. The recently emerged carbohydrate esterase (CE) 15 family of glucuronoyl esterases (GEs) has...

  2. Common and distant structural characteristics of feruloyl esterase families from Aspergillus oryzae.

    Directory of Open Access Journals (Sweden)

    D B R K Gupta Udatha

    Full Text Available BACKGROUND: Feruloyl esterases (FAEs are important biomass degrading accessory enzymes due to their capability of cleaving the ester links between hemicellulose and pectin to aromatic compounds of lignin, thus enhancing the accessibility of plant tissues to cellulolytic and hemicellulolytic enzymes. FAEs have gained increased attention in the area of biocatalytic transformations for the synthesis of value added compounds with medicinal and nutritional applications. Following the increasing attention on these enzymes, a novel descriptor based classification system has been proposed for FAEs resulting into 12 distinct families and pharmacophore models for three FAE sub-families have been developed. METHODOLOGY/PRINCIPAL FINDINGS: The feruloylome of Aspergillus oryzae contains 13 predicted FAEs belonging to six sub-families based on our recently developed descriptor-based classification system. The three-dimensional structures of the 13 FAEs were modeled for structural analysis of the feruloylome. The three genes coding for three enzymes, viz., A.O.2, A.O.8 and A.O.10 from the feruloylome of A. oryzae, representing sub-families with unknown functional features, were heterologously expressed in Pichia pastoris, characterized for substrate specificity and structural characterization through CD spectroscopy. Common feature-based pharamacophore models were developed according to substrate specificity characteristics of the three enzymes. The active site residues were identified for the three expressed FAEs by determining the titration curves of amino acid residues as a function of the pH by applying molecular simulations. CONCLUSIONS/SIGNIFICANCE: Our findings on the structure-function relationships and substrate specificity of the FAEs of A. oryzae will be instrumental for further understanding of the FAE families in the novel classification system. The developed pharmacophore models could be applied for virtual screening of compound databases for

  3. Common and distant structural characteristics of feruloyl esterase families from Aspergillus oryzae.

    Science.gov (United States)

    Udatha, D B R K Gupta; Mapelli, Valeria; Panagiotou, Gianni; Olsson, Lisbeth

    2012-01-01

    Feruloyl esterases (FAEs) are important biomass degrading accessory enzymes due to their capability of cleaving the ester links between hemicellulose and pectin to aromatic compounds of lignin, thus enhancing the accessibility of plant tissues to cellulolytic and hemicellulolytic enzymes. FAEs have gained increased attention in the area of biocatalytic transformations for the synthesis of value added compounds with medicinal and nutritional applications. Following the increasing attention on these enzymes, a novel descriptor based classification system has been proposed for FAEs resulting into 12 distinct families and pharmacophore models for three FAE sub-families have been developed. The feruloylome of Aspergillus oryzae contains 13 predicted FAEs belonging to six sub-families based on our recently developed descriptor-based classification system. The three-dimensional structures of the 13 FAEs were modeled for structural analysis of the feruloylome. The three genes coding for three enzymes, viz., A.O.2, A.O.8 and A.O.10 from the feruloylome of A. oryzae, representing sub-families with unknown functional features, were heterologously expressed in Pichia pastoris, characterized for substrate specificity and structural characterization through CD spectroscopy. Common feature-based pharamacophore models were developed according to substrate specificity characteristics of the three enzymes. The active site residues were identified for the three expressed FAEs by determining the titration curves of amino acid residues as a function of the pH by applying molecular simulations. Our findings on the structure-function relationships and substrate specificity of the FAEs of A. oryzae will be instrumental for further understanding of the FAE families in the novel classification system. The developed pharmacophore models could be applied for virtual screening of compound databases for short listing the putative substrates prior to docking studies or for post

  4. Contribution of soil esterase to biodegradation of aliphatic polyester agricultural mulch film in cultivated soils.

    Science.gov (United States)

    Yamamoto-Tamura, Kimiko; Hiradate, Syuntaro; Watanabe, Takashi; Koitabashi, Motoo; Sameshima-Yamashita, Yuka; Yarimizu, Tohru; Kitamoto, Hiroko

    2015-01-01

    The relationship between degradation speed of soil-buried biodegradable polyester film in a farmland and the characteristics of the predominant polyester-degrading soil microorganisms and enzymes were investigated to determine the BP-degrading ability of cultivated soils through characterization of the basal microbial activities and their transition in soils during BP film degradation. Degradation of poly(butylene succinate-co-adipate) (PBSA) film was evaluated in soil samples from different cultivated fields in Japan for 4 weeks. Both the degradation speed of the PBSA film and the esterase activity were found to be correlated with the ratio of colonies that produced clear zone on fungal minimum medium-agarose plate with emulsified PBSA to the total number colonies counted. Time-dependent change in viable counts of the PBSA-degrading fungi and esterase activities were monitored in soils where buried films showed the most and the least degree of degradation. During the degradation of PBSA film, the viable counts of the PBSA-degrading fungi and the esterase activities in soils, which adhered to the PBSA film, increased with time. The soil, where the film was degraded the fastest, recorded large PBSA-degrading fungal population and showed high esterase activity compared with the other soil samples throughout the incubation period. Meanwhile, esterase activity and viable counts of PBSA-degrading fungi were found to be stable in soils without PBSA film. These results suggest that the higher the distribution ratio of native PBSA-degrading fungi in the soil, the faster the film degradation is. This could be due to the rapid accumulation of secreted esterases in these soils.

  5. The effect of EDTA and metal cations on the 5-bromoindoxyl acetate esterase activity in the thyroid of the guinea pig

    DEFF Research Database (Denmark)

    Kirkeby, S

    1976-01-01

    Miscellaneous metal cations and EDTA have been used as activators and inhibitors of esterase activity in the thyroid of the guinea-pig. The results indicate that the 5-bromoiondoxyl acetate esterase in the epithelial cells probably consists of two different A-esterase isoenzymes, one present...

  6. Zymography Detection of a Bacterial Extracellular Thermoalkaline Esterase/Lipase Activity.

    Science.gov (United States)

    Tapizquent, María; Fernández, Marleny; Barreto, Georgina; Hernández, Zully; Contreras, Lellys M; Kurz, Liliana; Wilkesman, Jeff

    2017-01-01

    Lipases are esterases that occur widely in nature, yet those with commercial relevance are exclusively from microbial origin. Glycerol and long-chain fatty acids are the products after hydrolysis of esters bonds in saponifiable lipids catalyzed by lipases. In this work, we describe lipase/esterase activity contained in cell-free fractions from thermophilic bacteria, cultured in medium containing olive oil. Analysis of the cell-free fractions by electrotransference zymography, using tributyrin as substrate, revealed bands corresponding to lipase activity. The method is simple, fast, and inexpensive.

  7. Purification and properties of a novel quizalofop-p-ethyl-hydrolyzing esterase involved in quizalofop-p-ethyl degradation by Pseudomonas sp. J-2.

    Science.gov (United States)

    Zhang, Hui; Li, Mengya; Li, Jie; Wang, Guangli; Liu, Yuan

    2017-05-10

    Quizalofop-p-ethyl (QPE) is a post-emergence herbicide that effectively controls grass weeds and is often detected in the environment. However, the biochemical and molecular mechanisms of QPE degradation in the environment remains unclear. In this study, a highly effective QPE-degrading bacterial strain J-2 was isolated from acclimated activated sludge and identified as a Pseudomonas sp., containing the QPE breakdown metabolite quizalofop acid (QA) identified by Liquid Chromatography-Ion Trap-Mass Spectrometry (LC-IT-MS n ) analysis. A novel QPE hydrolase esterase-encoding gene qpeH was cloned from strain J-2 and functionally expressed in Escherichia coli BL21 (DE3). The specific activity of recombinant QpeH was 198.9 ± 2.7 U mg -1 for QPE with K m and K cat values of 41.3 ± 3.6 μM and 127.3 ± 4.5 s -1 . The optimal pH and temperature for the recombinant QpeH were 8.0 and 30 °C, respectively and the enzyme was activated by Ca 2+ , Cd 2+ , Li + , Fe 3+ and Co 2+ and inhibited by Ni 2+ , Fe 2+ , Ag + , DEPC, SDS, Tween 80, Triton X, β-mercaptoethanol, PMSF, and pCMB. In addition, the catalytic efficiency of QpeH toward different AOPP herbicides in descending order was as follows: fenoxaprop-P-ethyl > quizalofop-P-tefuryl > QPE > haloxyfop-P-methyl > cyhalofopbutyl > clodinafop-propargyl. On the basis of the phylogenetic analysis and multiple sequence alignment, the identified enzyme QpeH, was clustered with esterase family V, suggesting a new member of this family because of its low similarity of amino acid sequence with esterases reported previously.

  8. Mutations in the DNA-binding domain of NR2E3 affect in vivo dimerization and interaction with CRX.

    Directory of Open Access Journals (Sweden)

    Raphael Roduit

    Full Text Available BACKGROUND: NR2E3 (PNR is an orphan nuclear receptor essential for proper photoreceptor determination and differentiation. In humans, mutations in NR2E3 have been associated with the recessively inherited enhanced short wavelength sensitive (S- cone syndrome (ESCS and, more recently, with autosomal dominant retinitis pigmentosa (adRP. NR2E3 acts as a suppressor of the cone generation program in late mitotic retinal progenitor cells. In adult rod photoreceptors, NR2E3 represses cone-specific gene expression and acts in concert with the transcription factors CRX and NRL to activate rod-specific genes. NR2E3 and CRX have been shown to physically interact in vitro through their respective DNA-binding domains (DBD. The DBD also contributes to homo- and heterodimerization of nuclear receptors. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed NR2E3 homodimerization and NR2E3/CRX complex formation in an in vivo situation by Bioluminescence Resonance Energy Transfer (BRET(2. NR2E3 wild-type protein formed homodimers in transiently transfected HEK293T cells. NR2E3 homodimerization was impaired in presence of disease-causing mutations in the DBD, except for the p.R76Q and p.R104W mutant proteins. Strikingly, the adRP-linked p.G56R mutant protein interacted with CRX with a similar efficiency to that of NR2E3 wild-type and p.R311Q proteins. In contrast, all other NR2E3 DBD-mutant proteins did not interact with CRX. The p.G56R mutant protein was also more effective in abolishing the potentiation of rhodospin gene transactivation by the NR2E3 wild-type protein. In addition, the p.G56R mutant enhanced the transrepression of the M- and S-opsin promoter, while all other NR2E3 DBD-mutants did not. CONCLUSIONS/SIGNIFICANCE: These results suggest different disease mechanisms in adRP- and ESCS-patients carrying NR2E3 mutations. Titration of CRX by the p.G56R mutant protein acting as a repressor in trans may account for the severe clinical phenotype in adRP patients.

  9. E3 Testing of Directed Energy Systems: A Challenging Future

    National Research Council Canada - National Science Library

    Brown, Lyndell R

    2009-01-01

    .... Compatibility testing and susceptibility to electromagnetic radiation is required. Standards, such as MIL-STD-464 and MIL-STD-237D, are being revised to include HPM levels and frequencies for E3 tests...

  10. E3: Economy - Energy - Environment; Supporting Manufacturing Leadership through Sustainability

    Data.gov (United States)

    U.S. Environmental Protection Agency — The E3 initiative is designed to help you thrive in a new business era focused on sustainability and, working together, to promote sustainable manufacturing and...

  11. E3 Success Story - Working Together: E3 Ohio and the Ohio By-Product Synergy Network

    Science.gov (United States)

    The Mid-Ohio Regional Planning Commission (MORPC) received funding to support the integration of the national E3 sustainability initiative with the Ohio By-Product Synergy (BPS) Network to create an efficient and replicable model for reducing GHGs.

  12. Chaperone-like activities of α-synuclein: α-Synuclein assists enzyme activities of esterases

    International Nuclear Information System (INIS)

    Ahn, Misun; Kim, SeungBum; Kang, Mira; Ryu, Yeonwoo; Doohun Kim, T.

    2006-01-01

    α-Synuclein, a major constituent of Lewy bodies (LBs), has been implicated to play a critical role in the pathogenesis of Parkinson's disease (PD), although the physiological function of α-synuclein has not yet been known. Here we have shown that α-synuclein, which has no well-defined secondary or tertiary structure, can protect the enzyme activity of microbial esterases against stress conditions such as heat, pH, and organic solvents. In particular, the flexibility of α-synuclein and its C-terminal region seems to be important for complex formation, but the structural integrity of the C-terminal region may not be required for stabilization of enzyme activity. In addition, atomic force microscopy (AFM) and in vivo enzyme assays showed highly specific interactions of esterases with α-synuclein. Our results indicate that α-synuclein not only protects the enzyme activity of microbial esterases in vitro, but also can stabilize the active conformation of microbial esterases in vivo

  13. Esterase and protease activities of Bacillus spp. from afitin, iru and ...

    African Journals Online (AJOL)

    The electrophoretic profiles of fermented African locust bean protein (ALBP), using strains presenting the highest protease activities in casein agar, were analyzed by SDS-PAGE to select strains with good ability to be used as starter cultures. All the Bacillus spp. tested showed esterase activity against tributyrin with high ...

  14. Enrichment of maize and triticale bran with recombinant Aspergillus tubingensis ferulic acid esterase

    CSIR Research Space (South Africa)

    Zwane, EN

    2017-03-01

    Full Text Available acid, in particular for the enrichment of food substrates. A recombinant Aspergillus tubingensis ferulic acid esterase Type A (FAEA) was expressed in Aspergillus niger D15#26 and purified with anion-exchange chromatography (3487 U/mg, Km = 0.43 mM, Kcat...

  15. Lipase and esterase: to what extent can this classification be applied accurately?

    Directory of Open Access Journals (Sweden)

    Danielle Branta Lopes

    2011-09-01

    Full Text Available Enzyme technology is an ever-growing field of knowledge and, in recent years, this technology has raised renewed interest, due to the search for new paradigms in several productive processes. Lipases, esterases and cutinases are enzymes used in a wide range of processes involving synthesis and hydrolysis reactions. The objective of this work was to investigate and compare the specific lipase and esterase activities of five enzymes - four already classified as lipases and one classified as cutinase - in the presence of natural and synthetic substrates. All tested enzymes presented both esterase and lipase specific activities. The highest specific esterase activity was observed for Aspergillus 1068 lipase in natural substrate and for F. oxysporum cutinase in synthetic substrate, while the highest specific lipase activity was observed for Geotrichum sp. lipase in natural substrate and for F. oxysporum cutinase in synthetic substrate. These results display some interface-independent lipolytic activity for all lipases tested. This is in accordance with the rationale that a new and broader definition of lipases may be necessary.

  16. Distribution and substrate specificity of esterases in the housefly, Musca domestica L.

    NARCIS (Netherlands)

    Asperen, K. van

    1959-01-01

    Housefly homogenates perform high cholinesterase and ali-esterase activity. Warburg-manometric studies show that acetylcholine, acetyl-β-methylcholine, butyrylcholine, and benzoylcholine are exclusively hydrolysed by a cholinesterase, the properties of which are more or less comparable to those of

  17. Esterase isozymes patterns of grape vine (Vitis vinifera L. are altered in response to fungicide exposure

    Directory of Open Access Journals (Sweden)

    Gleice Ribeiro Orasmo

    2015-10-01

    Full Text Available Current analysis characterizes the effect of different fungicides often applied for pest control on a-and b-esterase patterns of four economically important table-wine grape cultivars (Italia, Rubi, Benitaka and Brasil of Vitis vinifera. The a- and b-esterase patterns in bud leaves of the cultivars were assessed by native PAGE analysis. Cabrio Top® compound inhibited Est-2, Est-5, Est-6, Est-7, Est-8, Est-9 and Est-10 carboxylesterases, whereas Est-4, Est-11, Est-12, Est-13, Est-14 acetylesterases and Est-16 carboxylesterase were detected as weakly stained bands. Carboxylesterases and acetylesterases were also detected as weakly stained bands when exposed to fungicides Orthocide 500®, Positron Duo® and Folicur PM®. No changes in a- and b-esterase patterns were reported when the vines were exposed to the fungicides Rovral SC®, Kumulus DF®, Curzate M®, Score® or Cuprogarb 500®. The evidence of functional changes in carboxylesterase and acetylesterase levels in current study is a warning to grape producers on the dangers inherent in the indiscriminate use of potent and modern fungicides extensively used in agriculture. The inhibition effect of fungicides on esterase isozyme molecules seems to be independent of the fungicide chemical.

  18. Fungal glucuronoyl esterases : Genome mining based enzyme discovery and biochemical characterization

    NARCIS (Netherlands)

    Dilokpimol, Adiphol; Mäkelä, Miia R; Cerullo, Gabriella; Zhou, Miaomiao; Varriale, Simona; Gidijala, Loknath; Brás, Joana L A; Jütten, Peter; Piechot, Alexander; Verhaert, Raymond; Faraco, Vincenza; Hilden, Kristiina S.; de Vries, Ronald P

    2018-01-01

    4-O-Methyl-d-glucuronic acid (MeGlcA) is a side-residue of glucuronoarabinoxylan and can form ester linkages to lignin, contributing significantly to the strength and rigidity of the plant cell wall. Glucuronoyl esterases (4-O-methyl-glucuronoyl methylesterases, GEs) can cleave this ester bond, and

  19. Cholesterol esterase inhibitory activity of bioactives from leaves of Mangifera indica L

    Science.gov (United States)

    Gururaja, G. M.; Mundkinajeddu, Deepak; Dethe, Shekhar M.; Sangli, Gopala K.; Abhilash, K.; Agarwal, Amit

    2015-01-01

    Background: In the earlier studies, methanolic extract of Mangifera indica L leaf was exhibited hypocholesterol activity. However, the bioactive compounds responsible for the same are not reported so far. Objective: To isolate the bioactive compounds with hypocholesterol activity from the leaf extract using cholesterol esterase inhibition assay which can be used for the standardization of extract. Materials and Methods: The leaf methanolic extract of M. indica (Sindoora variety) was partitioned with ethyl acetate and chromatographed on silica gel to yield twelve fractions and the activity was monitored by using cholesterol esterase inhibition assay. Active fractions were re-chromatographed to yield individual compounds. Results and Discussion: A major compound mangiferin present in the extract was screened along with other varieties of mango leaves for cholesterol esterase inhibition assay. However, the result indicates that compounds other than mangiferin may be active in the extract. Invitro pancreatic cholesterol esterase inhibition assay was used for bioactivity guided fractionation (BAGF) to yield bioactive compound for standardization of extract. Bioactivity guided fractionation afford the active fraction containing 3b-taraxerol with an IC50 value of 0.86μg/ml. Conclusion: This study demonstrates that M. indica methanol extract of leaf have significant hypocholesterol activity which is standardized with 3b-taraxerol, a standardized extract for hypocholesterol activity resulted in development of dietary supplement from leaves of Mangifera indica. PMID:26692750

  20. Dampak Hipoksia Sistemik terhadap Malondialdehida, Glial Fibrillary Acidic Protein dan Aktivitas Asetilkolin Esterase Otak Tikus

    OpenAIRE

    Andriani Andriani; Ani Retno Prijanti; Ninik Mudjihartini; Sri Widia A. Jusman

    2016-01-01

    Hipoksia sistemik menyebabkan berkurangnya oksigen dan energi di otak sehingga memicupenglepasan neurotransmiter asetilkolin, meningkatkan radikal bebas dan glial fibrillary acidic protein (GFAP)yang berfungsi menjaga kekuatan membran. Tujuan penelitian untuk melihat gambaran adaptasi otak padahipoksia sistemik terhadap fungsi asetilkolin esterase, kerusakan membran sel neuron dan astrosit. Penelitiandilakukan di Laboratorium Biokimia & Biologi Molekuler FK Universitas Indonesia, pada ta...

  1. Hydrolysis of Wheat Arabinoxylan by Two Acetyl Xylan Esterases from Chaetomium thermophilum

    DEFF Research Database (Denmark)

    Tong, Xiaoxue; Lange, Lene; Grell, Morten Nedergaard

    2015-01-01

    The thermophilic filamentous ascomycete Chaetomium thermophilum produces functionally diverse hemicellulases when grown on hemicellulose as carbon source. Acetyl xylan esterase (EC 3.1.1.72) is an important accessory enzyme in hemicellulose biodegradation. Although the genome of C. thermophilum has...

  2. Production and partial characterisation of feruloyl esterase by Sporotrichum thermophile in solid-state fermentation

    DEFF Research Database (Denmark)

    Topakas, E.; Kalogeris, E.; Kekos, D.

    2003-01-01

    A number of factors affecting production of feruloyl esterase an enzyme that hydrolyse ester linkages of ferulic acid (FA) in plant cell walls, by the thermophylic fungus Sporotrichum thermophile under solid state fermentation (SSF) were investigated. Initial moisture content and type of carbon...

  3. Usefulness of Leukocyte Esterase Test Versus Rapid Strep Test for Diagnosis of Acute Strep Pharyngitis

    Directory of Open Access Journals (Sweden)

    Kumara V. Nibhanipudi MD

    2015-08-01

    Full Text Available Objective: A study to compare the usage of throat swab testing for leukocyte esterase on a test strip(urine dip stick-multi stick to rapid strep test for rapid diagnosis of Group A Beta hemolytic streptococci in cases of acute pharyngitis in children. Hypothesis: The testing of throat swab for leukocyte esterase on test strip currently used for urine testing may be used to detect throat infection and might be as useful as rapid strep. Methods: All patients who come with a complaint of sore throat and fever were examined clinically for erythema of pharynx, tonsils and also for any exudates. Informed consent was obtained from the parents and assent from the subjects. 3 swabs were taken from pharyngo-tonsillar region, testing for culture, rapid strep & Leukocyte Esterase. Results: Total number is 100. Cultures 9(+; for rapid strep== 84(- and16 (+; For LE== 80(- and 20(+ Statistics: From data configuration Rapid Strep versus LE test don’t seem to be a random (independent assignment but extremely aligned. The Statistical results show rapid and LE show very agreeable results. Calculated Value of Chi Squared Exceeds Tabulated under 1 Degree Of Freedom (P<.0.0001 reject Null Hypothesis and Conclude Alternative Conclusions: Leukocyte esterase on throat swab is as useful as rapid strep test for rapid diagnosis of strep pharyngitis on test strip currently used for urine dip stick causing acute pharyngitis in children.

  4. Trichoderma reesei CE16 acetyl esterase and its role in enzymatic degradation of acetylated hemicellulose

    DEFF Research Database (Denmark)

    Biely, Peter; Cziszarava, Maria; Agger, Jane W.

    2014-01-01

    Results The combined action of GH10 xylanase and acetylxylan esterases (AcXEs) leads to formation of neutral and acidic xylooligosaccharides with a few resistant acetyl groups mainly at their non-reducing ends. We show here that these acetyl groups serve as targets for TrCE16 AcE. The most promin...

  5. Crystallization and preliminary X-ray diffraction studies of the pneumococcal teichoic acid phosphorylcholine esterase Pce

    Energy Technology Data Exchange (ETDEWEB)

    Lagartera, Laura; González, Ana; Stelter, Meike; García, Pedro; Kahn, Richard; Menéndez, Margarita; Hermoso, Juan A., E-mail: xjuan@iqfr.csic.es

    2005-02-01

    The modular choline-binding protein Pce, the phosphorylcholine esterase from S. pneumoniae, has been crystallized by the hanging-drop vapour-diffusion method. A SAD data set from a derivative with a gadolinium complex has been collected to 2.7 Å resolution.

  6. C1-esterase inhibitor blocks T lymphocyte proliferation and cytotoxic T lymphocyte generation in vitro

    DEFF Research Database (Denmark)

    Nissen, Mogens Holst; Bregenholt, S; Nording, J A

    1998-01-01

    We have previously shown that activated C1s complement and activated T cells cleave beta2-microglobulin (beta2m) in vitro leading to the formation of desLys58 beta2m. This process can specifically be inhibited by C1-esterase inhibitor (C1-inh). Furthermore we showed that exogenously added desLys58...

  7. Preliminary X-ray Study of Naproxen Esterase from Bacillus subtilis

    NARCIS (Netherlands)

    van der Laan, Jan; Teplyakov, A.V.; Lammers, A.A.; Dijkstra, B.W.

    1993-01-01

    Single crystals of naproxen esterase from Bacillus subtilis have been obtained from PEG6000 solutions at pH 8.0 by liquid-liquid diffusion while applying a temperature gradient from 4°C to room temperature over a period of four weeks. The crystals belong to the trigonal space group P3121 or P3221

  8. Non-specific esterases and esterproteases in masticatory muscles from the muscular dystrophic mouse

    DEFF Research Database (Denmark)

    Kirkeby, S; Moe, D; Vilmann, H

    1989-01-01

    With the aid of histochemical and electrophoretic techniques activities for esterase and esterprotease were investigated in the digastric and masseter muscles from normal and dystrophic mice. The substrates used were alpha-naphthyl acetate and N-acetyl-L-alanine alpha-naphthyl ester. According...

  9. A New Functional Classification of Glucuronoyl Esterases by Peptide Pattern Recognition

    DEFF Research Database (Denmark)

    Wittrup Agger, Jane; Busk, Peter Kamp; Pilgaard, Bo

    2017-01-01

    of characterized enzymes exist and the exact activity is still uncertain. Here peptide pattern recognition is used as a bioinformatic tool to identify and group new CE15 proteins that are likely to have glucuronoyl esterase activity. 1024 CE15-like sequences were drawn from GenBank and grouped into 24 groups...

  10. Crystal Structure and Substrate Specificity Modification of Acetyl Xylan Esterase from Aspergillus luchuensis.

    Science.gov (United States)

    Komiya, Dai; Hori, Akane; Ishida, Takuya; Igarashi, Kiyohiko; Samejima, Masahiro; Koseki, Takuya; Fushinobu, Shinya

    2017-10-15

    Acetyl xylan esterase (AXE) catalyzes the hydrolysis of the acetyl bonds present in plant cell wall polysaccharides. Here, we determined the crystal structure of AXE from Aspergillus luchuensis ( Al AXEA), providing the three-dimensional structure of an enzyme in the Esterase_phb family. Al AXEA shares its core α/β-hydrolase fold structure with esterases in other families, but it has an extended central β-sheet at both its ends and an extra loop. Structural comparison with a ferulic acid esterase (FAE) from Aspergillus niger indicated that Al AXEA has a conserved catalytic machinery: a catalytic triad (Ser119, His259, and Asp202) and an oxyanion hole (Cys40 and Ser120). Near the catalytic triad of A lAXEA, two aromatic residues (Tyr39 and Trp160) form small pockets at both sides. Homology models of fungal FAEs in the same Esterase_phb family have wide pockets at the corresponding sites because they have residues with smaller side chains (Pro, Ser, and Gly). Mutants with site-directed mutations at Tyr39 showed a substrate specificity similar to that of the wild-type enzyme, whereas those with mutations at Trp160 acquired an expanded substrate specificity. Interestingly, the Trp160 mutants acquired weak but significant type B-like FAE activity. Moreover, the engineered enzymes exhibited ferulic acid-releasing activity from wheat arabinoxylan. IMPORTANCE Hemicelluloses in the plant cell wall are often decorated by acetyl and ferulic acid groups. Therefore, complete and efficient degradation of plant polysaccharides requires the enzymes for cleaving the side chains of the polymer. Since the Esterase_phb family contains a wide array of fungal FAEs and AXEs from fungi and bacteria, our study will provide a structural basis for the molecular mechanism of these industrially relevant enzymes in biopolymer degradation. The structure of the Esterase_phb family also provides information for bacterial polyhydroxyalkanoate depolymerases that are involved in biodegradation of

  11. Novel E3 ubiquitin ligases that regulate histone protein levels in the budding yeast Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Rakesh Kumar Singh

    Full Text Available Core histone proteins are essential for packaging the genomic DNA into chromatin in all eukaryotes. Since multiple genes encode these histone proteins, there is potential for generating more histones than what is required for chromatin assembly. The positively charged histones have a very high affinity for negatively charged molecules such as DNA, and any excess of histone proteins results in deleterious effects on genomic stability and cell viability. Hence, histone levels are known to be tightly regulated via transcriptional, posttranscriptional and posttranslational mechanisms. We have previously elucidated the posttranslational regulation of histone protein levels by the ubiquitin-proteasome pathway involving the E2 ubiquitin conjugating enzymes Ubc4/5 and the HECT (Homologous to E6-AP C-Terminus domain containing E3 ligase Tom1 in the budding yeast. Here we report the identification of four additional E3 ligases containing the RING (Really Interesting New Gene finger domains that are involved in the ubiquitylation and subsequent degradation of excess histones in yeast. These E3 ligases are Pep5, Snt2 as well as two previously uncharacterized Open Reading Frames (ORFs YKR017C and YDR266C that we have named Hel1 and Hel2 (for Histone E3 Ligases respectively. Mutants lacking these E3 ligases are sensitive to histone overexpression as they fail to degrade excess histones and accumulate high levels of endogenous histones on histone chaperones. Co-immunoprecipitation assays showed that these E3 ligases interact with the major E2 enzyme Ubc4 that is involved in the degradation related ubiquitylation of histones. Using mutagenesis we further demonstrate that the RING domains of Hel1, Hel2 and Snt2 are required for histone regulation. Lastly, mutants corresponding to Hel1, Hel2 and Pep5 are sensitive to replication inhibitors. Overall, our results highlight the importance of posttranslational histone regulatory mechanisms that employ multiple E3

  12. Cloning, Expression and Characterization of a Thermostable Esterase HydS14 from Actinomadura sp. Strain S14 in Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Pichapak Sriyapai

    2015-06-01

    Full Text Available A thermostable esterase gene (hydS14 was cloned from an Actinomadura sp. S14 gene library. The gene is 777 bp in length and encodes a polypeptide of 258 amino acid residues with no signal peptide, no N-glycosylation site and a predicted molecular mass of 26,604 Da. The encoded protein contains the pentapeptide motif (GYSLG and catalytic triad (Ser88-Asp208-His235 of the esterase/lipase superfamily. The HydS14 sequence shows 46%–64% identity to 23 sequences from actinomycetes (23 α/β-hydrolases, has three conserved regions, and contains the novel motif (GY(FSLG, which distinguishes it from other clusters in the α/β-hydrolase structural superfamily. A plasmid containing the coding region (pPICZαA-hydS14 was used to express HydS14 in Pichia pastoris under the control of the AOXI promoter. The recombinant HydS14 collected from the supernatant had a molecular mass of ~30 kDa, which agrees with its predicted molecular mass without N-glycosylation. HydS14 had an optimum temperature of approximately 70 °C and an optimum pH of 8.0. HydS14 was stable at 50 and 60 °C for 120 min, with residual activities of above 80% and above 90%, respectively, as well as 50% activity at pH 6.0–8.0 and pH 9.0, respectively. The enzyme showed higher activity with p-nitrophenyl-C2 and C4. The Km and Vmax values for p-nitrophenyl-C4 were 0.21 ± 0.02 mM and 37.07 ± 1.04 μmol/min/mg, respectively. The enzyme was active toward short-chain p-nitrophenyl ester (C2–C6, displaying optimal activity with p-nitrophenyl-C4 (Kcat/Km = 11.74 mM−1·S−1. In summary, HydS14 is a thermostable esterase from Actinomadura sp. S14 that has been cloned and expressed for the first time in Pichia pastoris.

  13. Photochemical Aryl Radical Cyclizations to Give (E-3-Ylideneoxindoles

    Directory of Open Access Journals (Sweden)

    Michael Gurry

    2014-09-01

    Full Text Available (E-3-Ylideneoxindoles are prepared in methanol in reasonable to good yields, as adducts of photochemical 5-exo-trig of aryl radicals, in contrast to previously reported analogous radical cyclizations initiated by tris(trimethylsilylsilane and azo-initiators that gave reduced oxindole adducts.

  14. Apolipoprotein E*3-Leiden transgenic mice mode for hypolipidaemic drugs

    NARCIS (Netherlands)

    Vlijmen, B.J.M. van; Pearce, N.J.; Bergö, M.; Staels, B.; Yates, J.W.; Gribble, A.D.; Bond, B.C.; Hofker, M.H.; Havekes, L.M.; Groot, P.H.E.

    1998-01-01

    Apolipoprotein (APO) E*3-Leiden mice with impaired chylomicron and VLDL (very low density lipoprotein) remnant metabolism display hyperlipidaemia and atherosclerosis. In the present study, these mice were used for testing the hypolipidaemic effect of two marketed agents, lovastatin (CAS 75330-75-5)

  15. A carboxylesterase, Esterase-6, modulates sensory physiological and behavioral response dynamics to pheromone in Drosophila

    Directory of Open Access Journals (Sweden)

    Chertemps Thomas

    2012-06-01

    Full Text Available Abstract Background Insects respond to the spatial and temporal dynamics of a pheromone plume, which implies not only a strong response to 'odor on', but also to 'odor off'. This requires mechanisms geared toward a fast signal termination. Several mechanisms may contribute to signal termination, among which odorant-degrading enzymes. These enzymes putatively play a role in signal dynamics by a rapid inactivation of odorants in the vicinity of the sensory receptors, although direct in vivo experimental evidences are lacking. Here we verified the role of an extracellular carboxylesterase, esterase-6 (Est-6, in the sensory physiological and behavioral dynamics of Drosophila melanogaster response to its pheromone, cis-vaccenyl acetate (cVA. Est-6 was previously linked to post-mating effects in the reproductive system of females. As Est-6 is also known to hydrolyze cVA in vitro and is expressed in the main olfactory organ, the antenna, we tested here its role in olfaction as a putative odorant-degrading enzyme. Results We first confirm that Est-6 is highly expressed in olfactory sensilla, including cVA-sensitive sensilla, and we show that expression is likely associated with non-neuronal cells. Our electrophysiological approaches show that the dynamics of olfactory receptor neuron (ORN responses is strongly influenced by Est-6, as in Est-6° null mutants (lacking the Est-6 gene cVA-sensitive ORN showed increased firing rate and prolonged activity in response to cVA. Est-6° mutant males had a lower threshold of behavioral response to cVA, as revealed by the analysis of two cVA-induced behaviors. In particular, mutant males exhibited a strong decrease of male-male courtship, in association with a delay in courtship initiation. Conclusions Our study presents evidence that Est-6 plays a role in the physiological and behavioral dynamics of sex pheromone response in Drosophila males and supports a role of Est-6 as an odorant-degrading enzyme (ODE in male

  16. Direct Kinetic Evidence for the Formation of an Acylpyridinium Intermediate in Synthetic p-Nitrophenyl Esterase-Catalyzed Hydrolysis Reactions

    National Research Council Canada - National Science Library

    Wang, Guang-Jia

    1996-01-01

    .... The deacylation rate was also found to exhibit a maximum for the same substrate 2 (n=6). These results are similar to those previously reported with cholesterol esterase as catalyst for the same hydrolysis reaction...

  17. The natural catalytic function of CuGE glucuronoyl esterase in hydrolysis of genuine lignin-carbohydrate complexes from birch

    DEFF Research Database (Denmark)

    Mosbech, Caroline; Holck, Jesper; Meyer, Anne S.

    2018-01-01

    Glucuronoyl esterases belong to carbohydrate esterase family 15 and catalyze de-esterification. Their natural function is presumed to be cleavage of ester linkages in lignin-carbohydrate complexes particularly those linking lignin and glucuronoyl residues in xylans in hardwood. Here, we show...... for the first time a detailed product profile of aldouronic acids released from birchwood lignin by a glucuronoyl esterase from the white-rot fungus Cerrena unicolor (CuGE). CuGE releases substrate for GH10 endo-xylanase which results in significantly increased product release compared to the action of endo......-xylanase alone. CuGE also releases neutral xylo-oligosaccharides that can be ascribed to the enzymes feruloyl esterase side activity as demonstrated by release of ferulic acid from insoluble wheat arabinoxylan. The data verify the enzyme's unique ability to catalyze removal of all glucuronoxylan associated...

  18. Dampak Hipoksia Sistemik terhadap Malondialdehida, Glial Fibrillary Acidic Protein dan Aktivitas Asetilkolin Esterase Otak Tikus

    Directory of Open Access Journals (Sweden)

    Andriani Andriani

    2016-09-01

    Full Text Available Hipoksia sistemik menyebabkan berkurangnya oksigen dan energi di otak sehingga memicupenglepasan neurotransmiter asetilkolin, meningkatkan radikal bebas dan glial fibrillary acidic protein (GFAPyang berfungsi menjaga kekuatan membran. Tujuan penelitian untuk melihat gambaran adaptasi otak padahipoksia sistemik terhadap fungsi asetilkolin esterase, kerusakan membran sel neuron dan astrosit. Penelitiandilakukan di Laboratorium Biokimia & Biologi Molekuler FK Universitas Indonesia, pada tahun 2013.Penelitian ekperimental ini menggunakan hewan coba tikus spraque dawley yang diinduksi hipoksia sistemikyang diambil jaringan otak bagian korteks dan plasma tikus. Kelompok tikus terdiri atas kelompok kontrol,kelompok perlakuan induksi hipoksia hari ke-1, 3 hari, 5 hari dan hari ke-7. Parameter yang diukur adalahkadar malondialdehida (MDA otak dan plasma, aktivitas spesifik enzim AChE jaringan otak serta kadar GFAPjaringan otak. Hasil menunjukkan bahwa hipoksia sistemik tidak meningkatkankadar MDA otak dan plasma.Induksi hipoksia sistemik meningkatkan aktivitas spesifik enzim AChE dan kadar GFAP jaringan otak secarabermakna. Pada plasma tidak terjadi peningkatan kadar GFAP. Hipoksia sistemik selama hari ke-7 belummenyebabkan kerusakan oksidatif, namun memperlihatkan peningkatan aktivitas AChe dan adaptasi astrositmelalui peningkatan GFAP. Kata kunci: hipoksia, astrosit, glial fibrillary acidic protein, malondialdehida, asetilkolin esterase   Systemic Hypoxia Effect on Rat Brain Malondialdehyde, Glial FibrillaryAcidic Protein, and Acetylcholine Esterase Activity Abstract Sistemic hypoxia causes lack of oxygen and energy in brain that trigger the release of acetylcholine,free radical and Glial fibrillary acidic protein (GFAP, a specific protein in astrocyte cells that act to strenghtenastrocite membrane. The aim of the research was to evaluate the damages of brain in systemic hypoxiathrough activity of acetylcholine esterase, neuron and astrocyte membran

  19. Using a simple HPLC approach to identify the enzymatic products of UTL-5g, a small molecule TNF-α inhibitor, from porcine esterase and from rabbit esterase.

    Science.gov (United States)

    Swartz, Kenneth; Zhang, Yiguan; Valeriote, Frederick; Chen, Ben; Shaw, Jiajiu

    2013-12-01

    UTL-5g is a novel small-molecule chemoprotector that lowers hepatotoxicity, nephrotoxicity, and myelotoxicity induced by cisplatin through TNF-α inhibition among other factors. As a prelude to investigating the metabolites of UTL-5g, we set out to identify the enzymatic products of UTL-5g under the treatment of both porcine liver esterase (PLE) and rabbit liver esterase (RLE). First, a number of mixtures made by UTL-5g and PLE were incubated at 25°C. At predetermined time points, individual samples were quenched by acetonitrile, vortexed, and centrifuged. The supernatants were then analyzed by reversed-phase HPLC (using a C18 column). The retention times and UV/vis spectra of individual peaks were compared to those of UTL-5g and its two postulated enzymatic products; thus the enzymatic products of UTL-5g were tentatively identified. Secondly, a different HPLC method (providing different retentions times) was used to cross-check and to confirm the identities of the two enzymatic products. Based on the observations, it was concluded that under the treatment of PLE, the major enzymatic products of UTL-5g were 5-methyliosxazole-3-carboxylic acid (ISOX) and 2,4-dichloroaniline (DCA). Treatment of UTL-5g by RLE also provided the same enzymatic products of UTL-5g from esterase. These results indicate that the peptide bond in UTL-5g was cleaved by PLE/RLE. Michaelis-Menten kinetics showed that the Km values of UTL-5g were 2.07mM with PLE and 0.37mM with RLE indicating that UTL-5g had a higher affinity with RLE. In summary, by a simple HPLC approach, we have concluded that the peptide bond in UTL-5g was cleaved by esterase from either porcine liver or rabbit liver in vitro and afforded DCA (at a mole ratio of 1:1) and ISOX. However, further studies are needed in order to determine whether UTL-5g is metabolized by microsomal enzymes to produce ISOX and DCA. Copyright © 2013 Elsevier B.V. All rights reserved.

  20. Esterase activities of intracellular extracts of wild strains of lactic acid bacteria isolated from Serra da Estrela cheese

    OpenAIRE

    Macedo, Angela C.; Tavares, Tânia G.; Malcata, F. Xavier

    2003-01-01

    Lactococcus lactis subsp. lactis strain ESB110019 and Lactobacillus plantarum strain ESB5004, novel strains that were previously isolated from the wild adventitious microflora of certified Serra da Estrela cheeses, were assayed for esterase activity using, as substrates, ortho- and para-nitrophenyl derivatives of fatty acids. Both strains preferentially hydrolyzed short-chain fatty acids; L. lactis ESB110019 exhibited a stronger esterase activity than Lb. plantarum ESB5004 and cleaved the p-n...

  1. Third International Meeting on Esterases Reacting with Organophosphorus Compounds

    Science.gov (United States)

    1998-01-01

    cassette for negative selection, 884 bp of ACHE including exon 1, 1.6 kb of a Neor gene cassette for positive selection, 5.2 kb of the ACHE Bam HI...fragment including exon 6, and 3 kb of Bluescript. Deletion of exons 2-5 removed 80% of the ACHE coding sequence. The gene targeting vector was...expression due to environmental influences on CYP3A4 and the presence or absence of CYP3A5 which may be under genetic control in man. Plasma

  2. Potato tuber pectin structure is influenced by pectin methyl esterase activity and impacts on cooked potato texture.

    Science.gov (United States)

    Ross, Heather A; Wright, Kathryn M; McDougall, Gordon J; Roberts, Alison G; Chapman, Sean N; Morris, Wayne L; Hancock, Robert D; Stewart, Derek; Tucker, Gregory A; James, Euan K; Taylor, Mark A

    2011-01-01

    Although cooked potato tuber texture is an important trait that influences consumer preference, a detailed understanding of tuber textural properties at the molecular level is lacking. Previous work has identified tuber pectin methyl esterase activity (PME) as a potential factor impacting on textural properties. In this study, tuber PME isoform and gene expression profiles have been determined in potato germplasm with differing textural properties as assessed using an amended wedge fracture method and a sloughing assay, revealing major differences between the potato types. Differences in pectin structure between potato types with different textural properties were revealed using monoclonal antibodies specific for different pectic epitopes. Chemical analysis of tuber pectin clearly demonstrated that, in tubers containing a higher level of total PME activity, there was a reduced degree of methylation of cell wall pectin and consistently higher peak force and work done values during the fracture of cooked tuber samples, demonstrating the link between PME activity, the degree of methylation of cell wall pectin, and cooked tuber textural properties.

  3. Functional expression of an ajmaline pathway-specific esterase from Rauvolfia in a novel plant-virus expression system.

    Science.gov (United States)

    Ruppert, Martin; Woll, Jörn; Giritch, Anatoli; Genady, Ezzat; Ma, Xueyan; Stöckigt, Joachim

    2005-11-01

    Acetylajmalan esterase (AAE) plays an essential role in the late stage of ajmaline biosynthesis. Based on the partial peptide sequences of AAE isolated and purified from Rauvolfia cell suspensions, a full-length AAE cDNA clone was isolated. The amino acid sequence of AAE has the highest level of identity of 40% to putative lipases known from the Arabidopsis thaliana genome project. Based on the primary structure AAE is a new member of the GDSL lipase superfamily. The expression in Escherichia coli failed although a wide range of conditions were tested. With a novel virus-based plant expression system, it was possible to express AAE functionally in leaves of Nicotiana benthamiana Domin. An extraordinarily high enzyme activity was detected in the Nicotiana tissue, which exceeded that in Rauvolfia serpentina (L.) Benth. ex Kurz cell suspension cultures about 20-fold. This expression allowed molecular analysis of AAE for the first time and increased the number of functionally expressed alkaloid genes from Rauvolfia now to eight, and the number of ajmaline pathway-specific cDNAs to a total of six.

  4. An Aspergillus oryzae acetyl xylan esterase: molecular cloning and characteristics of recombinant enzyme expressed in Pichia pastoris.

    Science.gov (United States)

    Koseki, Takuya; Miwa, Yozo; Akao, Takeshi; Akita, Osamu; Hashizume, Katsumi

    2006-02-10

    We screened 20,000 clones of an expressed sequence tag (EST) library from Aspergillus oryzae (http://www.nrib.go.jp/ken/EST/db/index.html) and obtained one cDNA clone encoding a protein with similarity to fungal acetyl xylan esterase. We also cloned the corresponding gene, designated as Aoaxe, from the genomic DNA. The deduced amino acid sequence consisted of a putative signal peptide of 31-amino acids and a mature protein of 276-amino acids. We engineered Aoaxe for heterologous expression in P. pastoris. Recombinant AoAXE (rAoAXE) was secreted by the aid of fused alpha-factor secretion signal peptide and accumulated as an active enzyme in the culture medium to a final level of 190 mg/l after 5 days. Purified rAoAXEA before and after treatment with endoglycosidase H migrated by SDS-PAGE with a molecular mass of 31 and 30 kDa, respectively. Purified rAoAXE displayed the greatest hydrolytic activity toward alpha-naphthylacetate (C2), lower activity toward alpha-naphthylpropionate (C3) and no detectable activity toward acyl-chain substrates containing four or more carbon atoms. The recombinant enzyme catalyzed the release of acetic acid from birchwood xylan. No activity was detectable using methyl esters of ferulic, caffeic or sinapic acids. rAoAXE was thermolabile in comparison to other AXEs from Aspergillus.

  5. Properties of Two Novel Esterases Identified from Culture Supernatant of Penicillium purpurogenum Grown on Sugar Beet Pulp.

    Science.gov (United States)

    Oleas, Gabriela; Callegari, Eduardo; Sepulveda, Romina; Eyzaguirre, Jaime

    2016-01-01

    The filamentous fungus Penicillium purpurogenum grows on a variety of natural carbon sources, such as sugar beet pulp, and secretes to the medium a large number of enzymes that degrade the carbohydrate components of lignocellulose. Sugar beet pulp is rich in pectin, and the purpose of this work is to identify novel esterases produced by the fungus, which may participate in pectin degradation. Partially purified culture supernatants of the fungus grown on sugar beet pulp were subjected to mass spectrometry analysis. Peptides thus identified, which may be part of potential esterases were probed against the proteins deduced from the fungal genome sequence. The cDNAs of two putative esterases identified were expressed in Pichia pastoris and their properties studied. One of these enzymes, named FAET, is a feruloyl esterase, while the other, PE, is classified as a pectin methyl esterase. These findings add to our knowledge of the enzymology of pectin degradation by Penicillium purpurogenum, and define properties of two novel esterases acting on de-esterification of pectin. Their availability may be useful as tools for the study of pectin structure and degradation.

  6. Complete replication-competent adenovirus 11p vectors with E1 or E3 insertions show improved heat stability

    Energy Technology Data Exchange (ETDEWEB)

    Mei, Ya-Fang, E-mail: ya-fang.mei@umu.se [Department of Clinical Microbiology and Virology, Umeå University, SE-901 85 Umeå (Sweden); Wu, Haidong, E-mail: haidong.wu@umu.se [Department of Clinical Microbiology and Virology, Umeå University, SE-901 85 Umeå (Sweden); Hultenby, Kjell, E-mail: kjell.hultenby@ki.se [Division of Clinical Research Centre, Department of Laboratory Medicine, Karolinska Institute, SE-14186 Stockholm (Sweden); Silver, Jim, E-mail: jim.silver@umu.se [Department of Clinical Microbiology and Virology, Umeå University, SE-901 85 Umeå (Sweden)

    2016-10-15

    Conventional adenovirus vectors harboring E1 or E3 deletions followed by the insertion of an exogenous gene show considerably reduced virion stability. Here, we report strategies to generate complete replication-competent Ad11p(RCAd11p) vectors that overcome the above disadvantage. A GFP cassette was successfully introduced either upstream of E1A or in the E3A region. The resulting vectors showed high expression levels of the hexon and E1genes and also strongly induced the cytopathic effect in targeted cells. When harboring oversized genomes, the RCAd11pE1 and RCAd11pE3 vectors showed significantly improved heat stability in comparison to Ad11pwt;of the three, RCAd11pE3 was the most tolerant to heat treatment. Electron microscopy showed that RCAd11pE3, RCAd11pE1, Ad11pwt, and Ad11pE1 Delmanifested dominant, moderate, minimum, or no full virus particles after heat treatment at 47 °C for 5 h. Our results demonstrated that both genome size and the insertion site in the viral genome affect virion stability. -- Highlights: •Replicating adenovirus 11p GFP vectors at the E1 or E3 region were generated. •RCAd11pE3 and RCAd11pE1 vectors manifested significantly improved heat stability. •RCAd11pE3 and RCAd11pE1 showed more full viral particles than Ad11pwt after heating. •We demonstrated that both genome size and the insertion site affect virion stability.

  7. Complete replication-competent adenovirus 11p vectors with E1 or E3 insertions show improved heat stability

    International Nuclear Information System (INIS)

    Mei, Ya-Fang; Wu, Haidong; Hultenby, Kjell; Silver, Jim

    2016-01-01

    Conventional adenovirus vectors harboring E1 or E3 deletions followed by the insertion of an exogenous gene show considerably reduced virion stability. Here, we report strategies to generate complete replication-competent Ad11p(RCAd11p) vectors that overcome the above disadvantage. A GFP cassette was successfully introduced either upstream of E1A or in the E3A region. The resulting vectors showed high expression levels of the hexon and E1genes and also strongly induced the cytopathic effect in targeted cells. When harboring oversized genomes, the RCAd11pE1 and RCAd11pE3 vectors showed significantly improved heat stability in comparison to Ad11pwt;of the three, RCAd11pE3 was the most tolerant to heat treatment. Electron microscopy showed that RCAd11pE3, RCAd11pE1, Ad11pwt, and Ad11pE1 Delmanifested dominant, moderate, minimum, or no full virus particles after heat treatment at 47 °C for 5 h. Our results demonstrated that both genome size and the insertion site in the viral genome affect virion stability. -- Highlights: •Replicating adenovirus 11p GFP vectors at the E1 or E3 region were generated. •RCAd11pE3 and RCAd11pE1 vectors manifested significantly improved heat stability. •RCAd11pE3 and RCAd11pE1 showed more full viral particles than Ad11pwt after heating. •We demonstrated that both genome size and the insertion site affect virion stability.

  8. Chromatographic study of highly methoxylated lime pectins deesterified by different pectin methyl-esterases.

    Science.gov (United States)

    Ralet, M C; Bonnin, E; Thibault, J F

    2001-03-25

    The inter-molecular distribution of free carboxyl groups of two highly methoxylated pectins enzymatically deesterified by plant and fungus pectin methyl-esterases were investigated by size-exclusion (SEC) and ion-exchange chromatography (IEC). "Homogeneous" populations with respect to molar mass or charge density were thereby obtained and their chemical composition and physico-chemical properties (transport parameter for monovalent cations and calcium, calcium activity coefficient) were studied. Chemical analysis showed that the composition varies from one SEC fraction to another, the highest molar mass fraction being richer in rhamnose and galactose and exhibiting a slightly higher degree of methylation. Separation of pectins by IEC revealed a quite homogeneous charge density distribution for F58 contrary to P60 which exhibited a large distribution of methoxyl groups. The free carboxyl groups distributions and calcium binding behaviours of SEC and IEC fractions were shown to differ widely for highly methoxylated pectins deesterified by plant and fungus pectin methyl-esterases.

  9. Production and partial characterization of alkaline feruloyl esterases by Fusarium oxysporum during submerged batch cultivation

    DEFF Research Database (Denmark)

    Topakas, E.; Christakopoulos, Paul

    2004-01-01

    Production of feruloyl esterases (FAEs) by Fusarium oxysporum was enhanced by optimization of initial pH of the culture medium, the type and concentration of nitrogen and carbon source. Submerged batch cultivation in a laboratory bioreactor (17 1) produced activity at 82 nkat g(-1) dry substrate....... Production of FAE does not therefore, require FA, however, production is diminished by the removal of esterified FA from the growth substrate. Optimal FAE activity was observed at pH 7 and 50 degreesC with 68 and 55% activity at pH 8 and pH 9, respectively. The esterase was fully stable at pH 5-8 and up...

  10. Lunar Penetrating Radar onboard the Chang'e-3 mission

    Science.gov (United States)

    Fang, Guang-You; Zhou, Bin; Ji, Yi-Cai; Zhang, Qun-Ying; Shen, Shao-Xiang; Li, Yu-Xi; Guan, Hong-Fei; Tang, Chuan-Jun; Gao, Yun-Ze; Lu, Wei; Ye, Sheng-Bo; Han, Hai-Dong; Zheng, Jin; Wang, Shu-Zhi

    2014-12-01

    Lunar Penetrating Radar (LPR) is one of the important scientific instruments onboard the Chang'e-3 spacecraft. Its scientific goals are the mapping of lunar regolith and detection of subsurface geologic structures. This paper describes the goals of the mission, as well as the basic principles, design, composition and achievements of the LPR. Finally, experiments on a glacier and the lunar surface are analyzed.

  11. Energy Exascale Earth System Model (E3SM) Project Strategy

    Energy Technology Data Exchange (ETDEWEB)

    Bader, D. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2017-09-18

    The E3SM project will assert and maintain an international scientific leadership position in the development of Earth system and climate models at the leading edge of scientific knowledge and computational capabilities. With its collaborators, it will demonstrate its leadership by using these models to achieve the goal of designing, executing, and analyzing climate and Earth system simulations that address the most critical scientific questions for the nation and DOE.

  12. Characterization of a cold-adapted esterase and mutants from a psychotolerant Pseudomonas sp. strain.

    Science.gov (United States)

    Dong, Juan; Gasmalla, Mohammed A A; Zhao, Wei; Sun, Jingtao; Liu, Wenyu; Wang, Mingming; Han, Liang; Yang, Ruijin

    2017-09-01

    A cold-adapted esterase-producing strain named T1-39 was isolated from Glacier No. 1, Tianshan, People's Republic of China and identified as Pseudomonas sp. from 16S rRNA sequence analysis. The esterase (EstT1-39) secreted by this strain preferentially hydrolyzed esters of glycerol with short- and medium-chain fatty acids. Mutants of T1-39 were generated by the atmospheric and room temperature plasma method and screened for enhanced esterase activity. Among all the mutants, strain TB11 had 4.45-fold higher esterase productivity than T1-39, with high genetic stability over 10 generations of continuous cultivation. Maximum activity of EstT1-39 and EstTB11 was observed at 30 ℃, pH 9.0 and 25 ℃, pH 8.5, respectively. EstTB11 was thermally more stable (50 ℃ for 1 H) and active over a broader pH range than EstT1-39. EstTB11 also retained 38% of its maximal activity at 0 ℃ and was found to be able to hydrolyze milk fats into short- and medium-chain fatty acids at 4 ℃. The characteristics of EstT1-39 made it a cold-adapted enzyme and the EstTB11 from the mutant, with its higher activity at lower temperatures, may be suitable for the production of aromas and flavors in the dairy industry. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

  13. Environmental Factors Modulating the Stability and Enzymatic Activity of the Petrotoga mobilis Esterase (PmEst.

    Directory of Open Access Journals (Sweden)

    Jose L S Lopes

    Full Text Available Enzymes isolated from thermophilic organisms found in oil reservoirs can find applications in many fields, including the oleochemical, pharmaceutical, bioenergy, and food/dairy industries. In this study, in silico identification and recombinant production of an esterase from the extremophile bacteria Petrotoga mobilis (designated PmEst were performed. Then biochemical, bioinformatics and structural characterizations were undertaken using a combination of synchrotron radiation circular dichroism (SRCD and fluorescence spectroscopies to correlate PmEst stability and hydrolytic activity on different substrates. The enzyme presented a high Michaelis-Menten constant (KM 0.16 mM and optimum activity at ~55°C for p-nitrophenyl butyrate. The secondary structure of PmEst was preserved at acid pH, but not under alkaline conditions. PmEst was unfolded at high concentrations of urea or guanidine through apparently different mechanisms. The esterase activity of PmEst was preserved in the presence of ethanol or propanol and its melting temperature increased ~8°C in the presence of these organic solvents. PmEst is a mesophilic esterase with substrate preference towards short-to medium-length acyl chains. The SRCD data of PmEst is in agreement with the prediction of an α/β protein, which leads us to assume that it displays a typical fold of esterases from this family. The increased enzyme stability in organic solvents may enable novel applications for its use in synthetic biology. Taken together, our results demonstrate features of the PmEst enzyme that indicate it may be suitable for applications in industrial processes, particularly, when the use of polar organic solvents is required.

  14. EVALUATION OF ESTERASE POLYMORPHISMS IN MATURE SEEDS OF RADISH (RAPHANUS SATIVUS L. ACCESSIONS OF VIR COLLECTION

    Directory of Open Access Journals (Sweden)

    A. S. Rudakova

    2017-01-01

    Full Text Available A biochemical evaluation of 25 radish accessions (Raphanus sativus L. on esterase isozymes of mature seeds has been carried out. The results of the experiments showed a wide range of diversity among the genotypes based on electrophoretic zones of esterase isoenzymes. The revealed isoenzyme complex of esterases was represented by eight isoforms with molecular weights from 37.7 kD to 57.6 kD. All accessions were divided into 13 electrophoretic zymotypes, differing from each other by the presence or absence of definite zones. The most often observed electrophoretic zymo-type is Gr. 1, which includes 24% of the total number of accessions evaluated. There are 8 zymotypes (Gr. 6 Gr. 13 with a frequency of occurrence 4%. Three groups (Gr. 2 – Gr. 4 had the same frequency of occurrence – 12%. Zimotype of Gr. 5 containes the maximum number of zones – 8. 2 zimotypes – Gr. 3 and Gr. 12 had the smallest number of 4 zones. Two zones of esterases – zones 7 and 8 (Мr 39.7кD and Мr 37.7 kD, respectively were monomorphic. The remaining six zones were polymorphic, i.e. could be absent in some zimotypes. The frequency of occurrence of each zone in different zymotypes has varied from 6.58% to 17.11%. As results of this research the accessions that were selected can become the most promising parent forms for future genetic and selection studies of this culture.

  15. Common and Distant Structural Characteristics of Feruloyl Esterase Families from Aspergillus oryzae

    OpenAIRE

    Udatha, D. B. R. K. Gupta; Mapelli, Valeria; Panagiotou, Gianni; Olsson, Lisbeth

    2012-01-01

    Background: Feruloyl esterases (FAEs) are important biomass degrading accessory enzymes due to their capability of cleaving the ester links between hemicellulose and pectin to aromatic compounds of lignin, thus enhancing the accessibility of plant tissues to cellulolytic and hemicellulolytic enzymes. FAEs have gained increased attention in the area of biocatalytic transformations for the synthesis of value added compounds with medicinal and nutritional applications. Following the increasing a...

  16. Interactions between resin monomers and commercial composite resins with human saliva derived esterases.

    Science.gov (United States)

    Jaffer, F; Finer, Y; Santerre, J P

    2002-04-01

    Cholesterol esterase (CE) and pseudocholinesterase (PCE) have been reported to degrade commercial and model composite resins containing bisphenylglycidyl dimethacrylate (BisGMA), triethylene glycol dimethacrylate (TEGDMA) or the latter in combination with urethane modified BisGMA monomer systems. In addition, human saliva has been shown to contain esterase like activities similar to CE and PCE. Hence, it was the aim of the current study to determine to what extent human saliva could degrade two common commercial composite resins (Z250 from 3M Inc. and Spectrum TPH from L.D. Caulk) which contain the above monomer systems. Saliva samples from different volunteers were collected, processed, pooled, and freeze-dried. TEGDMA and BisGMA monomers were incubated with human saliva derived esterase activity (HSDEA) and their respective hydrolysis was monitored using high performance liquid chromatography (HPLC). Both monomers were completely hydrolyzed within 25 h by HSDEA. Photopolymerized composites were incubated with buffer or human saliva (pH 7.0 and 37 C) for 2, 8 and 16 days. The incubation solutions were analyzed using HPLC and mass spectrometry. Surface morphology characterization was carried out using scanning electron microscopy. Upon biodegradation, the Z250 composite yielded higher amounts of BisGMA and TEGDMA related products relative to the TPH composite. However, there were higher amounts of ethoxylated bis-phenol A released from the TPH material. In terms of total mass of products released, human saliva demonstrated a greater ability to degrade Z250. In summary, HSDEA has been shown to contain esterase activities that can readily catalyze the biodegradation of current commercial composite resins.

  17. Hepatic esterase activity is increased in hepatocyte-like cells derived from human embryonic stem cells using a 3D culture system.

    Science.gov (United States)

    Choi, Young-Jun; Kim, Hyemin; Kim, Ji-Woo; Yoon, Seokjoo; Park, Han-Jin

    2018-05-01

    The aim of the study is to generate a spherical three-dimensional (3D) aggregate of hepatocyte-like cells (HLCs) differentiated from human embryonic stem cells and to investigate the effect of the 3D environment on hepatic maturation and drug metabolism. Quantitative real-time PCR analysis indicated that gene expression of mature hepatocyte markers, drug-metabolizing enzymes, and hepatic transporters was significantly higher in HLCs cultured in the 3D system than in those cultured in a two-dimensional system (p formation, were increased in HLCs cultured in the 3D system. In particular, 3D spheroidal culture increased expression of CES1 and BCHE, which encode hepatic esterases (p 3D spheroidal culture enhances the maturation and drug metabolism of stem cell-derived HLCs, and this may help to optimize hepatic differentiation protocols for hepatotoxicity testing.

  18. A New Esterase from Thermobifida halotolerans Hydrolyses Polyethylene Terephthalate (PET and Polylactic Acid (PLA

    Directory of Open Access Journals (Sweden)

    Georg Steinkellner

    2012-02-01

    Full Text Available A new esterase from Thermobifida halotolerans (Thh_Est was cloned and expressed in E. coli and investigated for surface hydrolysis of polylactic acid (PLA and polyethylene terephthalate (PET. Thh_Est is a member of the serine hydrolases superfamily containing the -GxSxG- motif with 85–87% homology to an esterase from T. alba, to an acetylxylan esterase from T. fusca and to various Thermobifida cutinases. Thh_Est hydrolyzed the PET model substrate bis(benzoyloxyethylterephthalate and PET releasing terephthalic acid and mono-(2-hydroxyethyl terephthalate in comparable amounts (19.8 and 21.5 mmol/mol of enzyme while no higher oligomers like bis-(2-hydroxyethyl terephthalate were detected. Similarly, PLA was hydrolyzed as indicated by the release of lactic acid. Enzymatic surface hydrolysis of PET and PLA led to a strong hydrophilicity increase, as quantified with a WCA decrease from 90.8° and 75.5° to 50.4° and to a complete spread of the water drop on the surface, respectively.

  19. Purification and biochemical characterization of feruloyl esterases from Aspergillus terreus MTCC 11096.

    Science.gov (United States)

    Kumar, C Ganesh; Kamle, Avijeet; Kamal, Ahmed

    2013-01-01

    Aspergillus terreus MTCC 11096 isolated from the soils of agricultural fields cultivating sweet sorghum was previously identified to produce feruloyl esterases (FAEs). The enzymes responsible for feruloyl esterase activity were purified to homogeneity and named as AtFAE-1, AtFAE-2, and AtFAE-3. The enzymes were monomeric having molecular masses of 74, 23 and 36 kDa, respectively. Active protein bands were identified by a developed pH-dependent zymogram on native PAGE. The three enzymes exhibited variation in pH tolerance ranging between pH 5-8 and thermostability of up to 55°C. Inhibition studies revealed that the serine residue was essential for feruloyl esterase activity; moreover aspartyl and glutamyl residues are not totally involved at the active site. Metal ions such as Ca(2+), K(+), and Mg(2+) stabilized the enzyme activity for all three FAEs. Kinetic data indicated that all three enzymes showed catalytic efficiencies (k(cat) /K(m)) against different synthesized alkyl and aryl esters indicating their broad substrate specificity. The peptide mass fingerprinting by MALDI/TOF-MS analysis and enzyme affinity toward methoxy and hydroxy substituents on the benzene ring revealed that the AtFAE-1 belonged to type A while AtFAE-2 and AtFAE-3 were type C FAE. The FAEs could release 65 to 90% of ferulic acid from agrowaste substrates in the presence of xylanase. © 2013 American Institute of Chemical Engineers.

  20. Mechanism-Guided Discovery of an Esterase Scaffold with Promiscuous Amidase Activity

    Directory of Open Access Journals (Sweden)

    Charlotte Kürten

    2016-06-01

    Full Text Available The discovery and generation of biocatalysts with extended catalytic versatilities are of immense relevance in both chemistry and biotechnology. An enhanced atomistic understanding of enzyme promiscuity, a mechanism through which living systems acquire novel catalytic functions and specificities by evolution, would thus be of central interest. Using esterase-catalyzed amide bond hydrolysis as a model system, we pursued a simplistic in silico discovery program aiming for the identification of enzymes with an internal backbone hydrogen bond acceptor that could act as a reaction specificity shifter in hydrolytic enzymes. Focusing on stabilization of the rate limiting transition state of nitrogen inversion, our mechanism-guided approach predicted that the acyl hydrolase patatin of the α/β phospholipase fold would display reaction promiscuity. Experimental analysis confirmed previously unknown high amidase over esterase activity displayed by the first described esterase machinery with a protein backbone hydrogen bond acceptor to the reacting NH-group of amides. The present work highlights the importance of a fundamental understanding of enzymatic reactions and its potential for predicting enzyme scaffolds displaying alternative chemistries amenable to further evolution by enzyme engineering.

  1. Discovery of potential cholesterol esterase inhibitors using in silico docking studies

    Directory of Open Access Journals (Sweden)

    Thirumalaisamy Sivashanmugam

    2013-08-01

    Full Text Available New drug discovery is considered broadly in terms of two kinds of investiga-tional activities such as exploration and exploitation. This study deals with the evaluation of the cholesterol esterase inhibitory activity of flavonoids apigenin, biochanin, curcumin, diosmetin, epipervilline, glycitein, okanin, rhamnazin and tangeritin using in silico docking studies. In silico docking studies were carried out using AutoDock 4.2, based on the Lamarckian genetic algorithm principle. The results showed that all the selected flavonoids showed binding energy ranging between -7.08 kcal/mol to -5.64 kcal/mol when compared with that of the standard compound gallic acid (-4.11 kcal/mol. Intermolecular energy (-9.13 kcal/mol to -7.09 kcal/mol and inhibition constant (6.48 µM to 73.18 µM of the ligands also coincide with the binding energy. All the selected flavonoids contributed cholesterol esterase inhibitory activity, these molecular docking analyses could lead to the further develop-ment of potent cholesterol esterase inhibitors for the treatment of obesity.

  2. Eco-friendly surface modification on polyester fabrics by esterase treatment

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Jindan; Cai, Guoqiang; Liu, Jinqiang; Ge, Huayun; Wang, Jiping, E-mail: jipingwanghz@gmail.com

    2014-03-01

    Graphical abstract: - Highlights: • We used a simple and easy way to measure the enzyme activity. • We studied the mechanism by characterizing the chemical changes in the surface of fabric. • We studied the advantages in surface wettability, fiber integrity and mechanical performance of cutinase treated fabrics. • Cutinase pretreated fibers exhibited much improved fabric wicking and better fiber integrity comparing to alkali treated ones. • Cutinase pretreatment technology promotes energy conservation and emission reduction. - Abstract: Currently, traditional alkali deweighting technology is widely used to improve the hydrophilicity of polyester fabrics. However, the wastewater and heavy chemicals in the effluent cause enormous damage to the environment. Esterase treatment, which is feasible in mild conditions with high selectivity, can provide a clean and efficient way for polyester modification. Under the optimum conditions, the polyester fabric hydrolysis process of esterase had a linear kinetics. X-ray photoelectron spectrometry (XPS) results showed that hydroxyl and carboxyl groups were produced only on the surface of modified fiber without changing the chemical composition of the bulk. These fibers exhibited much improved fabric wicking, as well as greatly improved oily stain removal performance. Compared to the harsh alkali hydrolysis, the enzyme treatment led to smaller weight loss and better fiber integrity. The esterase treatment technology is promising to produce higher-quality polyester textiles with an environmental friendly approach.

  3. Some Preliminary Scientific Results of Chang'E-3 Mission

    Science.gov (United States)

    Zou, Y.; Li, W.; Zheng, Y.; Li, H.

    2015-12-01

    Chang'E-3 mission is the main task of Phase two of China Lunar Exploration Program (CLEP), and also is Chinese first probe of landing, working and roving on the moon. Chang'E-3 craft composed of a lander and a rover, and each of them carry four scientific payloads respectively. The landing site of Chang'E-3 was located at 44.12 degrees north latitude and 19.51 degrees west longitude, where is in the northern part of Imbrium Which the distance in its west direction from the landing site of former Soviet probe Luna-17 is about 400 km, and about 780km far from the landing site of Appolo-17 in its southeast direction. Unfortunately, after a series of scientific tests and exploration on the surface of the moon, the motor controller communication of the rover emerged a breakdown on January 16, 2014, which leaded the four payloads onboard the rover can't obtain data anymore. However, we have received some interesting scientific data which have been studied by Chinese scientists. During the landing process of Chang'E-3, the Landing camera got total 4673 images with the Resolution in millimeters to meters, and the lander and rover took pictures for each other at different point with Topography camera and Panoramic camera. We can find characteristic changes in celestial brightness with time by analyzing image data from Lunar-based Ultraviolet Telescope (LUT) and an unprecedented constraint on water content in the sunlit lunar exosphere seen by LUT). The figure observed by EUV camera (EUVC) shows that there is a transient weak area of the Earth's plasma sphere; This event took place about three hours. The scientists think that it might be related to the change of the particle density of mid-latitude ionosphere. The preliminary spectral and mineralogical results from the landing site are derived according to the data of Visible and Near-infrared Imaging Spectrometer (VNIS). Seven major elements including Mg, Al, Si, K, Ca, Ti and Fe have been identified by the Active Particle

  4. E3D, 3-D Elastic Seismic Wave Propagation Code

    International Nuclear Information System (INIS)

    Larsen, S.; Harris, D.; Schultz, C.; Maddix, D.; Bakowsky, T.; Bent, L.

    2004-01-01

    1 - Description of program or function: E3D is capable of simulating seismic wave propagation in a 3D heterogeneous earth. Seismic waves are initiated by earthquake, explosive, and/or other sources. These waves propagate through a 3D geologic model, and are simulated as synthetic seismograms or other graphical output. 2 - Methods: The software simulates wave propagation by solving the elasto-dynamic formulation of the full wave equation on a staggered grid. The solution scheme is 4-order accurate in space, 2-order accurate in time

  5. Aspects of radiative K{sup +}{sub e3} decays

    Energy Technology Data Exchange (ETDEWEB)

    Kubis, B. [Universitaet Bonn, Helmholtz-Institut fuer Strahlen- und Kernphysik, Bonn (Germany); Mueller, E.H. [Universitaet Bonn, Helmholtz-Institut fuer Strahlen- und Kernphysik, Bonn (Germany); University of Edinburgh, School of Physics, Edinburgh (United Kingdom); Gasser, J.; Schmid, M. [Universitaet Bern, Institut fuer theoretische Physik, Bern (Switzerland)

    2007-04-15

    We re-investigate the radiative charged kaon decay K{sup {+-}}{yields}{pi}{sup 0}e{sup {+-}}{nu}{sub e}{gamma} [K{sub e3{gamma}}{sup {+-}}] in chiral perturbation theory, merging the chiral expansion with Low's theorem. We thoroughly analyze the precision of the predicted branching ratio relative to the non-radiative decay channel. Structure dependent terms and their impact on differential decay distributions are investigated in detail, and the possibility to see effects of the chiral anomaly in this decay channel is emphasized. (orig.)

  6. Aspects of radiative K+e3 decays

    International Nuclear Information System (INIS)

    Kubis, B.; Mueller, E.H.; Gasser, J.; Schmid, M.

    2007-01-01

    We re-investigate the radiative charged kaon decay K ± →π 0 e ± ν e γ [K e3γ ± ] in chiral perturbation theory, merging the chiral expansion with Low's theorem. We thoroughly analyze the precision of the predicted branching ratio relative to the non-radiative decay channel. Structure dependent terms and their impact on differential decay distributions are investigated in detail, and the possibility to see effects of the chiral anomaly in this decay channel is emphasized. (orig.)

  7. ISG15 inhibits Nedd4 ubiquitin E3 activity and enhances the innate antiviral response.

    Science.gov (United States)

    Malakhova, Oxana A; Zhang, Dong-Er

    2008-04-04

    Interferons regulate diverse immune functions through the transcriptional activation of hundreds of genes involved in anti-viral responses. The interferon-inducible ubiquitin-like protein ISG15 is expressed in cells in response to a variety of stress conditions like viral or bacterial infection and is present in its free form or is conjugated to cellular proteins. In addition, protein ubiquitination plays a regulatory role in the immune system. Many viruses modulate the ubiquitin (Ub) pathway to alter cellular signaling and the antiviral response. Ubiquitination of retroviral group-specific antigen precursors and matrix proteins of the Ebola, vesicular stomatitis, and rabies viruses by Nedd4 family HECT domain E3 ligases is an important step in facilitating viral release. We found that Nedd4 is negatively regulated by ISG15. Free ISG15 specifically bound to Nedd4 and blocked its interaction with Ub-E2 molecules, thus preventing further Ub transfer from E2 to E3. Furthermore, overexpression of ISG15 diminished the ability of Nedd4 to ubiquitinate viral matrix proteins and led to a decrease in the release of Ebola VP40 virus-like particles from the cells. These results point to a mechanistically novel function of ISG15 in the enhancement of the innate anti-viral response through specific inhibition of Nedd4 Ub-E3 activity. To our knowledge, this is the first example of a Ub-like protein with the ability to interfere with Ub-E2 and E3 interaction to inhibit protein ubiquitination.

  8. Gene

    Data.gov (United States)

    U.S. Department of Health & Human Services — Gene integrates information from a wide range of species. A record may include nomenclature, Reference Sequences (RefSeqs), maps, pathways, variations, phenotypes,...

  9. E3-transitions in sup(105, 107, 109, 111)Ag

    International Nuclear Information System (INIS)

    Shevelev, G.A.; Troitskaya, A.G.; Kartashov, V.M.

    1978-01-01

    Electron radiation of the isomeric transitions of the sup(105-111)Ag odd nuclei was studied using an iron magnetic πsup(√2) beta spectrometer. For most isomeric transitions, relative intensities of the K, L, M, and N lines have been measured; for sup(105-111)Ag and 111 Cd they were measured for the first time. Energy of gamma transitions, relative intensities of internal conversion electrons (ICE) compared with the theoretical ICE values for the E3 transitions are presented. The observations for all the shells are in a fairly gool agreement with the calculations. Systematics of low-lying excited states of the silver nuclei involved is proposed. It has been established that spins and parities of the first excited states of the sup(105-111)Ag odd nuclei are 7/2 + . Multipolarities of isomeric transitions from these staes are pure E3. Spin and parity 9/2 + of the second excited states may be uniquely determined unly for 109 Ag from direct measurements of the ICE transition at 45.8 keV

  10. Implication of SUMO E3 ligases in nucleotide excision repair.

    Science.gov (United States)

    Tsuge, Maasa; Kaneoka, Hidenori; Masuda, Yusuke; Ito, Hiroki; Miyake, Katsuhide; Iijima, Shinji

    2015-08-01

    Post-translational modifications alter protein function to mediate complex hierarchical regulatory processes that are crucial to eukaryotic cellular function. The small ubiquitin-like modifier (SUMO) is an important post-translational modification that affects transcriptional regulation, nuclear localization, and the maintenance of genome stability. Nucleotide excision repair (NER) is a very versatile DNA repair system that is essential for protection against ultraviolet (UV) irradiation. The deficiencies in NER function remarkably increase the risk of skin cancer. Recent studies have shown that several NER factors are SUMOylated, which influences repair efficiency. However, how SUMOylation modulates NER has not yet been elucidated. In the present study, we performed RNAi knockdown of SUMO E3 ligases and found that, in addition to PIASy, the polycomb protein Pc2 affected the repair of cyclobutane pyrimidine dimers. PIAS1 affected both the removal of 6-4 pyrimidine pyrimidone photoproducts and cyclobutane pyrimidine dimers, whereas other SUMO E3 ligases did not affect the removal of either UV lesion.

  11. Trajectory Determination for Chang 'e-3 Probe Soft-landing

    Science.gov (United States)

    Yezhi, S.; Huang, Y.

    2017-12-01

    On December 2, 2013, The Chang 'e-3 (ce-3) probe was successfully launched from a long march-3b carrier rocket at Xichang satellite launch center. After more than five days of flying, the probe was captured by the moon to 100 km by 100 km. The orbit maneuvered to 15 km by 100 km 4 days later. Finally, at 21:12 Beijing time on December 14, 2013, it landed at the junction of the Sinus Iridum and Mare Imbrium. In the ce-3 project, the combined test mode of the radio ranging measurement and very long baseline interferometry (VLBI) was used. The soft-landing was carried out in ce-3 mission for the sampling .The paper presents a new method of trajectory determination for soft landing and sampling returning for lunar probe by B spline approximation. By simulation and data processing of Chang'E-3(CE-3), it could be assumed that the accuracy of trajectory determination of soft landing is less than 100 meters in CE-3. It appears that the difference between the endpoint of trajectory and the location from image processed by NASA'S LRO is less than 50m .It confirms the method of soft landing trajectory determination provided by the paper is effective. The paper analyzes the dynamics and control characteristics of the sampling returning, provides the preliminary feasible trajectory determination method for soft landing and sampling return of Chang'E-5 (CE - 5).

  12. E3 analysis for crude and vacuum distillation system

    Energy Technology Data Exchange (ETDEWEB)

    Mittal, V; Zhang, J; Yang, X; Xu, Q [Dan F. Smith Department of Chemical Engineering, Lamar University, Beaumont, Texas (United States)

    2011-11-15

    Crude oil blending is a very common practice in petroleum refineries, where the main focus is to minimize the total purchase cost of crude oils under specified blending oil properties. Crude oil blending actually has significant impacts on energy consumption from heating furnaces during crude oil processing. Conceivably, furnace energy consumption from burning fuels such as natural gas, fuel oil, or propane causes huge amounts of CO{sub 2} emissions. In this paper, a methodology framework for crude oil blending and processing with simultaneous consideration of energy, emission, and economic profit (E3) is developed. It includes four stages of work: steady-state modeling, heating energy consumption calculation, emission model development, and economic evaluation. With Aspen HYSYS simulation, the developed methodology provides a quantitative support for refinery to identify an optimal E3 operating strategy. A case study is implemented to demonstrate the efficacy of this methodology. (Copyright copyright 2011 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  13. Performance Measurement of Advanced Stirling Convertors (ASC-E3)

    Science.gov (United States)

    Oriti, Salvatore M.

    2013-01-01

    NASA Glenn Research Center (GRC) has been supporting development of the Advanced Stirling Radioisotope Generator (ASRG) since 2006. A key element of the ASRG project is providing life, reliability, and performance testing data of the Advanced Stirling Convertor (ASC). The latest version of the ASC (ASC-E3, to represent the third cycle of engineering model test hardware) is of a design identical to the forthcoming flight convertors. For this generation of hardware, a joint Sunpower and GRC effort was initiated to improve and standardize the test support hardware. After this effort was completed, the first pair of ASC-E3 units was produced by Sunpower and then delivered to GRC in December 2012. GRC has begun operation of these units. This process included performance verification, which examined the data from various tests to validate the convertor performance to the product specification. Other tests included detailed performance mapping that encompassed the wide range of operating conditions that will exist during a mission. These convertors were then transferred to Lockheed Martin for controller checkout testing. The results of this latest convertor performance verification activity are summarized here.

  14. The E3 ubiquitin ligase RNF185 facilitates the cGAS-mediated innate immune response.

    Directory of Open Access Journals (Sweden)

    Qiang Wang

    2017-03-01

    Full Text Available The cyclic GMP-AMP synthase (cGAS, upon cytosolic DNA stimulation, catalyzes the formation of the second messenger 2'3'-cGAMP, which then binds to stimulator of interferon genes (STING and activates downstream signaling. It remains to be elucidated how the cGAS enzymatic activity is modulated dynamically. Here, we reported that the ER ubiquitin ligase RNF185 interacted with cGAS during HSV-1 infection. Ectopic-expression or knockdown of RNF185 respectively enhanced or impaired the IRF3-responsive gene expression. Mechanistically, RNF185 specifically catalyzed the K27-linked poly-ubiquitination of cGAS, which promoted its enzymatic activity. Additionally, Systemic Lupus Erythematosus (SLE patients displayed elevated expression of RNF185 mRNA. Collectively, this study uncovers RNF185 as the first E3 ubiquitin ligase of cGAS, shedding light on the regulation of cGAS activity in innate immune responses.

  15. Molecular population genetics of the β-esterase gene cluster of ...

    Indian Academy of Sciences (India)

    Unknown

    The lines within each population are grouped according to their genetic similarity over the full sequence length. ..... (P = 0.048); Var ≥ 0.00094 (P = 0.012); modified Var ≥. 0.00207 ..... phila as well as in a variety of organisms (Balakirev and.

  16. Thermophilic acetylxylan esterase genes and enzymes from alicyclobacillus acidocaldarius and related organisms and methods

    Science.gov (United States)

    Thompson, Vicki S.; Thompson, David N.; Reed, David W.; Lacey, Jeffrey A.; Apel, William A.

    2018-01-09

    A genetically modified organism comprising at least one nucleic acid sequence and/or at least one recombinant nucleic acid isolated from Alicyclobacillus acidocaldarius and encoding a polypeptide involved in at least partially degrading, cleaving, transporting, metabolizing, or removing polysaccharide, lignocellulose, hemicellulose, lignin, chitin, heteroxylan, and/or xylan-decorating group; and at least one nucleic acid sequence and/or at least one recombinant nucleic acid encoding a polypeptide involved in fermenting sugar molecules to a product. Additionally, enzymatic and/or proteinaceous extracts may be isolated from one or more genetically modified organisms. The extracts are utilized to convert biomass into a product. Further provided are methods of converting biomass into products comprising: placing the genetically modified organism and/or enzymatic extracts thereof in fluid contact with polysaccharides, cellulose, lignocellulose, hemicellulose, lignin, starch, sugars, sugar oligomers, carbohydrates, complex carbohydrates, chitin, heteroxylans, glycosides, and/or xylan-, glucan-, galactan-, or mannan-decorating groups.

  17. The effect of C1-esterase inhibitor in definite and suspected streptococcal toxic shock syndrome. Report of seven patients.

    Science.gov (United States)

    Fronhoffs, S; Luyken, J; Steuer, K; Hansis, M; Vetter, H; Walger, P

    2000-10-01

    To evaluate the effect of adjunctive C1-esterase inhibitor substitution therapy on clinical characteristics and outcome of patients with streptococcal toxic shock syndrome (TSS). Observational. Medizinische Poliklinik, University of Bonn, Germany. Seven patients with direct or indirect evidence of streptococcal TSS. In addition to conventional and supportive therapy, all patients received 2-3 single doses of C1-esterase inhibitor totaling 6,000-10,000 U within the first 24 h after admission. All patients developed fulminant septic shock, multiorgan failure and/or capillary leak syndrome and necrotizing fasciitis within 10-72 h following the onset of first symptoms. Between 1 and 4 days following administration of C1-esterase inhibitor, a marked shift of fluid from extravascular to intravascular compartments took place in all but one patient, accompanied by a transient intra-alveolar lung edema and rapidly decreasing need for adrenergic agents. Six of seven patients survived. These clinical observations in a small series of patients and the favorable outcome point towards a positive effect of early and high-dose administration of C1-esterase inhibitor as adjunctive therapy in streptococcal TSS. The possible mechanism involved may be the attenuation of capillary leak syndrome (CLS) via early inactivation of complement and contact systems. Controlled studies are needed to establish an improvement of the survival rates of patients with streptococcal TSS following administration of C1-esterase inhibitor.

  18. Radiative K{sub e3} decays revisited

    Energy Technology Data Exchange (ETDEWEB)

    Gasser, J. [Universitaet Bern, Institut fuer Theoretische Physik, Bern (Switzerland); Kubis, B. [Universitaet Bern, Institut fuer Theoretische Physik, Bern (Switzerland); Universitaet Bonn, Helmholtz-Institut fuer Strahlen- und Kernphysik, Bonn (Germany); Paver, N. [Universita degli Studi di Trieste, Dipartimento di Fisica Teorica, Trieste (Italy); INFN-Trieste, Trieste (Italy); Verbeni, M. [Universidad de Granada, Departamento de Fisica Teorica y del Cosmos, Granada (Spain)

    2005-03-01

    Motivated by recent experimental results and ongoing measurements, we review the chiral perturbation theory prediction for K{sub L}{yields}{pi}{sup -+}e{sup {+-}}{nu}{sub e}{gamma} decays. Special emphasis is given to the stability of the inner bremsstrahlung-dominated relative branching ratio versus the K{sub e3} form factors, and on the separation of the structure-dependent amplitude in differential distributions over the phase space. For the structure-dependent terms, an assessment of the order p{sup 6} corrections is given, in particular, a full next-to-leading order calculation of the axial component is performed. The experimental analysis of the photon energy spectrum is discussed, and other potentially useful distributions are introduced. (orig.)

  19. The EISCAT_3D Project in Norway: E3DN

    Science.gov (United States)

    La Hoz, C.; Oksavik, K.

    2013-12-01

    EISCAT_3D (E3D) is a project to build the next generation of incoherent scatter radars endowed with 3-dimensional scalar and vector capabilities that will replace the current EISCAT radars in Northern Scandinavia. One active (transmitting) site in Norway and four passive (receiving) sites in the Nordic countries will provide 3-D vector imaging capabilities by rapid scanning and multi-beam forming. The unprecedented flexibility of the solid-state transmitter with high duty-cycle, arbitrary wave-forming and polarisation and its pulsed power of 10 MW will provide unrivalled experimental capabilities to investigate the highly non-stationary and non-homogeneous state of the polar upper atmosphere. Aperture Synthesis Imaging Radar (ASIR) will to endow E3D with imaging capabilities in 3-dimensions that includes sub-beam resolution. Complemented by pulse compression, it will provide 3-dimensional images of certain types of incoherent scatter radar targets resolved to about 100 metres at 100 km range, depending on the signal-to-noise ratio. The Norwegian scientific programme is inspired by the pioneer polar scientist Kristian Birkeland (picture) and includes pressing questions on polar upper atmospheric research, among others: (Q1) How to proceed beyond the present simplistic, static, stationary and homogeneous analysis of upper atmospheric and ionospheric processes? (Q2) How does space weather affect ionospheric processes and how to support modelling and space weather services? (Q3) How to advance fundamental plasma physics by employing the ionosphere as a natural plasma physics laboratory? (Q4) How does the influx of extraterrestrial material interact with the upper atmosphere and where does the material originate from? (Q5) How does solar activity couple from geospace into the lower atmosphere and climate system, and does this energy change the wave forcing of geospace from below? Kristian Birkeland, Norwegian scientist and pioneer in polar and auroral research.

  20. Phylogenetic analysis of the SINA/SIAH ubiquitin E3 ligase family in Metazoa.

    Science.gov (United States)

    Pepper, Ian J; Van Sciver, Robert E; Tang, Amy H

    2017-08-07

    The RAS signaling pathway is a pivotal developmental pathway that controls many fundamental biological processes including cell proliferation, differentiation, movement and apoptosis. Drosophila Seven-IN-Absentia (SINA) is a ubiquitin E3 ligase that is the most downstream signaling "gatekeeper" whose biological activity is essential for proper RAS signal transduction. Vertebrate SINA homologs (SIAHs) share a high degree of amino acid identity with that of Drosophila SINA. SINA/SIAH is the most conserved signaling component in the canonical EGFR/RAS/RAF/MAPK signal transduction pathway. Vertebrate SIAH1, 2, and 3 are the three orthologs to invertebrate SINA protein. SINA and SIAH1 orthologs are found in all major taxa of metazoans. These proteins have four conserved functional domains, known as RING (Really Interesting New Gene), SZF (SIAH-type zinc finger), SBS (substrate binding site) and DIMER (Dimerization). In addition to the siah1 gene, most vertebrates encode two additional siah genes (siah2 and siah3) in their genomes. Vertebrate SIAH2 has a highly divergent and extended N-terminal sequence, while its RING, SZF, SBS and DIMER domains maintain high amino acid identity/similarity to that of SIAH1. But unlike vertebrate SIAH1 and SIAH2, SIAH3 lacks a functional RING domain, suggesting that SIAH3 may be an inactive E3 ligase. The SIAH3 subtree exhibits a high degree of amino acid divergence when compared to the SIAH1 and SIAH2 subtrees. We find that SIAH1 and SIAH2 are expressed in all human epithelial cell lines examined thus far, while SIAH3 is only expressed in a limited subset of cancer cell lines. Through phylogenetic analyses of metazoan SINA and SIAH E3 ligases, we identified many invariant and divergent amino acid residues, as well as the evolutionarily conserved functional motifs in this medically relevant gene family. Our phylomedicinal study of this unique metazoan SINA/SIAH protein family has provided invaluable evolution-based support towards future

  1. Altered gene regulation and potential association with metabolic resistance development to imidacloprid in the tarnished plant bug, Lygus lineolaris.

    Science.gov (United States)

    Zhu, Yu Cheng; Luttrell, Randall

    2015-01-01

    Chemical spray on cotton is almost an exclusive method for controlling tarnished plant bug (TPB), Lygus lineolaris. Frequent use of imidacloprid is a concern for neonicotinoid resistance in this key pest. Information of how and why TPB becomes less susceptible to imidacloprid is essential for effective monitoring and managing resistance. Microarray analysis of 6688 genes in imidacloprid-selected TPB (Im1500FF) revealed 955 upregulated and 1277 downregulated (≥twofold) genes in Im1500FF, with 369 and 485 of them annotated. Five P450 and nine esterase genes were significantly upregulated, and only one esterase gene and no P450 genes were downregulated. Other upregulated genes include helicases, phosphodiesterases, ATPases and kinases. Pathway analyses identified 65 upregulated cDNAs that encode 51 different enzymes involved in 62 different pathways, including P450 and esterase genes for drug and xenobiotic metabolisms. Sixty-four downregulated cDNAs code only 17 enzymes that are associated with only 23 pathways mostly related to food digestion. This study demonstrated a significant change in gene expression related to metabolic processes in imidacloprid-selected TPB, resulting in overexpression of P450 and esterase genes for potential excess detoxification and cross/multiple resistance development. The identification of these and other enzyme genes establishes a foundation to explore the complicity of potential imidacloprid resistance in TPB. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

  2. Histochemical studies on genetical control of hormonal enzyme inducibility in the mouse. IV: Cellular localization of androgen sensitive nonspecific esterase in the epididymis

    DEFF Research Database (Denmark)

    Kirkeby, S; Blecher, S R

    1981-01-01

    Nonspecific esterase of mouse epididymis has previously been studied histochemically, using alpha naphthyl-acetate and 5-bromoindoxyl acetate techniques, as well as certain inhibitors. Epithelial cell types of the epididymis have been characterized, and certain esterase isozymes in a particular...

  3. Crystallization and preliminary X-ray diffraction analysis of the glucuronoyl esterase catalytic domain from Hypocrea jecorina

    International Nuclear Information System (INIS)

    Wood, S. J.; Li, X.-L.; Cotta, M. A.; Biely, P.; Duke, N. E. C.; Schiffer, M.; Pokkuluri, P. R.

    2008-01-01

    The catalytic domain of the glucuronoyl esterase from H. jecorina was overexpresssed, purified and crystallized in space group P2 1 2 1 2 1 . X-ray diffraction data were collected to 1.9 Å resolution. The catalytic domain of the glucuronoyl esterase from Hypocrea jecorina (anamorph Trichoderma reesei) was overexpresssed, purified and crystallized by the sitting-drop vapor-diffusion method using 1.4 M sodium/potassium phosphate pH 6.9. The crystals belonged to space group P2 1 2 1 2 1 and X-ray diffraction data were collected to 1.9 Å resolution. This is the first enzyme with glucoronoyl esterase activity to be crystallized; its structure will be valuable in lignocellulose-degradation research

  4. Crystallization and preliminary X-ray diffraction analysis of the glucuronoyl esterase catalytic domain from Hypocrea jecorina

    Energy Technology Data Exchange (ETDEWEB)

    Wood, S. J. [Biosciences Division, Argonne National Laboratory, Argonne, IL 60439 (United States); Li, X.-L.; Cotta, M. A. [Fermentation Biotechnology Research Unit, National Center for Agricultural Utilization Research, USDA-ARS, Peoria, Illinois 61604 (United States); Biely, P. [Institute of Chemistry, Slovak Academy of Sciences, 845 38 Bratislava (Slovakia); Duke, N. E. C.; Schiffer, M.; Pokkuluri, P. R., E-mail: rajp@anl.gov [Biosciences Division, Argonne National Laboratory, Argonne, IL 60439 (United States)

    2008-04-01

    The catalytic domain of the glucuronoyl esterase from H. jecorina was overexpresssed, purified and crystallized in space group P2{sub 1}2{sub 1}2{sub 1}. X-ray diffraction data were collected to 1.9 Å resolution. The catalytic domain of the glucuronoyl esterase from Hypocrea jecorina (anamorph Trichoderma reesei) was overexpresssed, purified and crystallized by the sitting-drop vapor-diffusion method using 1.4 M sodium/potassium phosphate pH 6.9. The crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1} and X-ray diffraction data were collected to 1.9 Å resolution. This is the first enzyme with glucoronoyl esterase activity to be crystallized; its structure will be valuable in lignocellulose-degradation research.

  5. Crystallization and preliminary crystallographic analysis of Axe2, an acetylxylan esterase from Geobacillus stearothermophilus

    International Nuclear Information System (INIS)

    Lansky, Shifra; Alalouf, Onit; Solomon, Vered; Alhassid, Anat; Govada, Lata; Chayan, Naomi E.; Belrhali, Hassan; Shoham, Yuval; Shoham, Gil

    2013-01-01

    The serine acetylxylan esterase from G. stearothermophilus (Axe2) has been crystallized in the tetragonal space group I422. Complete diffraction data sets have been measured for the selenomethionine derivative (SAD data, 1.70 Å resolution) and the wild-type enzyme (1.85 Å resolution) to be used for a full three-dimensional structural analysis of the Axe2 protein. Acetylxylan esterases are part of the hemi-cellulolytic system of many microorganisms which utilize plant biomass for growth. Xylans, which are polymeric sugars that constitute a significant part of the plant biomass, are usually substituted with acetyl side groups attached at position 2 or 3 of the xylose backbone units. Acetylxylan esterases hydrolyse the ester linkages of the xylan acetyl groups and thus improve the ability of main-chain hydrolysing enzymes to break down the sugar backbone units. As such, these enzymes play an important part in the hemi-cellulolytic utilization system of many microorganisms that use plant biomass for growth. Interest in the biochemical characterization and structural analysis of these enzymes stems from their numerous potential biotechnological applications. An acetylxylan esterase (Axe2) of this type from Geobacillus stearothermophilus T-6 has recently been cloned, overexpressed, purified, biochemically characterized and crystallized. One of the crystal forms obtained (RB1) belonged to the tetragonal space group I422, with unit-cell parameters a = b = 110.2, c = 213.1 Å. A full diffraction data set was collected to 1.85 Å resolution from flash-cooled crystals of the wild-type enzyme at 100 K using synchrotron radiation. A selenomethionine derivative of Axe2 has also been prepared and crystallized for single-wavelength anomalous diffraction experiments. The crystals of the selenomethionine-derivatized Axe2 appeared to be isomorphous to those of the wild-type enzyme and enabled the measurement of a full 1.85 Å resolution diffraction data set at the selenium

  6. Molecular characterization of a novel family VIII esterase from burkholderia multivorans UWC10

    CSIR Research Space (South Africa)

    Rashamuse, KJ

    2007-02-01

    Full Text Available et al., 1992). Here we report the cloning, purification, and 3D model of a novel family VIII esterase from Burkholderia multivorans UWC10. To our knowledge no report of esterolytic activity from B. multivorans is currently available. METHODOLOGY... stream_source_info Rashamuse1_2007_d.pdf.txt stream_content_type text/plain stream_size 9884 Content-Encoding UTF-8 stream_name Rashamuse1_2007_d.pdf.txt Content-Type text/plain; charset=UTF-8 Molecular Characterization...

  7. Esterase activity able to hydrolyze dietary antioxidant hydroxycinnamates is distributed along the intestine of mammals

    DEFF Research Database (Denmark)

    Andreasen, Mette Findal; Kroon, P A; Williamson, G

    2001-01-01

    and may contribute to the beneficial effects derived from consumption of cereal bran. However, these compounds are ester linked to the main polymers in the plant cell wall and cannot be absorbed in this complex form. The present work shows that esterases with activity toward esters of the major dietary......Hydroxycinnamic acids are effective antioxidants and are abundant components of plant cell walls, especially in cereal bran. For example, wheat and rye brans are rich sources of the hydroxycinnamates ferulic acid, sinapic acid, and p-coumaric acid. These phenolics are part of human and animal diets...

  8. Isolation and characterization of a novel cold-adapted esterase, MtEst45, from Microbulbifer thermotolerans DAU221

    Directory of Open Access Journals (Sweden)

    Yong-Suk eLee

    2016-03-01

    Full Text Available A novel esterase, MtEst45, was isolated from a fosmid genomic library of Microbulbifer thermotolerans DAU221. The encoding gene is predicted to have a mass of 45,564 Da and encodes 495 amino acids, excluding a 21 amino acid signal peptide. MtEst45 showed a low amino acid identity (approximately 23–24% compared with other lipolytic enzymes belonging to Family III, a closely related bacterial lipolytic enzyme family. MtEst45 also showed a conserved GXSXG motif, G131IS133YG135, which was reported as active site of known lipolytic enzymes, and the putative catalytic triad composed of D237 and H265. Because these mutants of MtEst45, which was S133A, D237N, and H265L, had no activity, these catalytic triad essential for the enzyme catalysis. MtEst45 was overexpressed in Escherichia coli BL21 (DE3 and purified via His-tag affinity chromatography. The optimal pH and temperature of MtEst45 were estimated to be 8.17 and 46.27°C by response surface methodology, respectively. Additionally, MtEst45 was also active between 1–15°C. The optimal hydrolysis substrate for MtEst45 among p-nitrophenyl esters (C2–C18 was p-nitrophenyl butyrate, and the Km and Vmax values were 0.0998 mM and 550 μmol/min/mg of protein, respectively. MtEst45 was strongly inhibited by Hg2+, Zn2+, and Cu2+ ions; by phenylmethanesulfonyl fluoride; and by β-mercaptoethanol. Ca2+ did not affect the enzyme’s activity. These biochemical properties, sequence identity, and phylogenetic analysis suggest that MtEst45 represents a novel and valuable bacterial lipolytic enzyme family and is useful for biotechnological applications.

  9. The effect of increased yeast alcohol acetyltransferase and esterase activity on the flavour profiles of wine and distillates.

    Science.gov (United States)

    Lilly, Mariska; Bauer, Florian F; Lambrechts, Marius G; Swiegers, Jan H; Cozzolino, Daniel; Pretorius, Isak S

    2006-07-15

    The fruity odours of wine are largely derived from the synthesis of esters and higher alcohols during yeast fermentation. The ATF1- and ATF2-encoded alcohol acetyltransferases of S. cerevisiae are responsible for the synthesis of ethyl acetate and isoamyl acetate esters, while the EHT1-encoded ethanol hexanoyl transferase is responsible for synthesizing ethyl caproate. However, esters such as these might be degraded by the IAH1-encoded esterase. The objectives of this study were: (a) to overexpress the genes encoding ester-synthesizing and ester-degrading enzymes in wine yeast; (b) to prepare Colombard table wines and base wines for distillation using these modified strains; and (c) to analyse and compare the ester concentrations and aroma profiles of these wines and distillates. The overexpression of ATF1 significantly increased the concentrations of ethyl acetate, isoamyl acetate, 2-phenylethyl acetate and ethyl caproate, while the overexpression of ATF2 affected the concentrations of ethyl acetate and isoamyl acetate to a lesser degree. The overexpression of IAH1 resulted in a significant decrease in ethyl acetate, isoamyl acetate, hexyl acetate and 2-phenylethyl acetate. The overexpression of EHT1 resulted in a marked increase in ethyl caproate, ethyl caprylate and ethyl caprate. The flavour profile of the wines and distillates prepared using the modified strains were also significantly altered as indicated by formal sensory analysis. This study offers prospects for the development of wine yeast starter strains with optimized ester-producing capability that could assist winemakers in their effort to consistently produce wine and distillates such as brandy to definable flavour specifications and styles.

  10. Fibulin-1C, C1 esterase inhibitor and glucose regulated protein 75 interact with the CREC proteins, calumenin and reticulocalbin

    DEFF Research Database (Denmark)

    Hansen, Gry Aune Westergaard; Ludvigsen, Maja; Jacobsen, Christian

    2015-01-01

    Affinity purification, immunoprecipitation, gel electrophoresis and mass spectrometry were used to identify fibulin-1C, C1 esterase inhibitor and glucose regulated protein 75, grp75, as binding partners of the CREC proteins, calumenin and reticulocalbin. Surface plasmon resonance was used to verify...... the interaction of all three proteins with each of the CREC proteins. Fibulin-1C interacts with calumenin and reticulocalbin with an estimated dissociation constant around 50-60 nM. The interaction, at least for reticulocalbin, was not dependent upon the presence of Ca2+. C1 esterase inhibitor interacted...

  11. Arginine-esterase activity of kallikrein in the sera of whole-body irradiated rats and guinea-pigs

    Energy Technology Data Exchange (ETDEWEB)

    Pouckova, P; Pospisil, J; Dienstbier, Z [Karlova Univ., Prague (Czechoslovakia). Biofyzikalni Ustav

    1977-09-01

    In whole-body irradiated rats (800 R=LDsub(50/30)) and guinea pigs (300 R=LDsub(50/30)) changes were investigated in the arginine esterase activity of kallikrein in native serum as well as in serum exposed to contact with a clay suspension. From the values obtained the activity of prekallikrein was calculated. While in the rat serum significant changes in the arginine esterase activity of kallikrein were found, in the guinea pig serum the kallikrein activity did not change markedly. The activity of prekallikrein immediately after irradiation assumes a similar course in both types of laboratory animals while during later intervals a reverse pattern was observed.

  12. Payload topography camera of Chang'e-3

    International Nuclear Information System (INIS)

    Yu, Guo-Bin; Liu, En-Hai; Zhao, Ru-Jin; Zhong, Jie; Zhou, Xiang-Dong; Zhou, Wu-Lin; Wang, Jin; Chen, Yuan-Pei; Hao, Yong-Jie

    2015-01-01

    Chang'e-3 was China's first soft-landing lunar probe that achieved a successful roving exploration on the Moon. A topography camera functioning as the lander's “eye” was one of the main scientific payloads installed on the lander. It was composed of a camera probe, an electronic component that performed image compression, and a cable assembly. Its exploration mission was to obtain optical images of the lunar topography in the landing zone for investigation and research. It also observed rover movement on the lunar surface and finished taking pictures of the lander and rover. After starting up successfully, the topography camera obtained static images and video of rover movement from different directions, 360° panoramic pictures of the lunar surface around the lander from multiple angles, and numerous pictures of the Earth. All images of the rover, lunar surface, and the Earth were clear, and those of the Chinese national flag were recorded in true color. This paper describes the exploration mission, system design, working principle, quality assessment of image compression, and color correction of the topography camera. Finally, test results from the lunar surface are provided to serve as a reference for scientific data processing and application. (paper)

  13. Characterisation of esterases as potential biomarkers of pesticide exposure in the lugworm Arenicola marina (Annelida: Polychaeta)

    International Nuclear Information System (INIS)

    Hannam, Marie L.; Hagger, Josephine A.; Jones, Malcolm B.; Galloway, Tamara S.

    2008-01-01

    Here, we identify and characterise cholinesterase (ChE) and carboxylesterase (CbE) activities in the body tissues of the sediment dwelling worm Arenicola marina. Exposure to the organophosphorus pesticide azamethiphos yielded an in vitro IC 50 of 5 μg l -1 for propionylcholinesterase (PChE). PChE was significantly inhibited in vivo after a 10 day exposure to 100 μg l -1 azamethiphos, equivalent to the recommended aquatic application rate (ANOVA; F = 2.75, P = 0.033). To determine sensitivity to environmental conditions, A. marina were exposed for 10 days to field collected sediments. PChE activity was significantly lower in worms exposed to sediments from an estuary classified to be at high risk from point source pollution by the UK Environment Agency (ANOVA; F = 15.33, P < 0.001). Whilst causality cannot be directly attributed from these latter exposures, they provide an important illustration of the potential utility of esterase activity as a biomarker of environmental quality in this ecologically relevant sentinel species. - This paper provides a preliminary characterisation of esterase enzyme activities in the tissues and body fluids of the sediment dwelling worm Arenicola marina and explores their potential use as biomarkers of organophosphorus pesticide exposure in the marine environment

  14. Modelling substrate specificity and enantioselectivity for lipases and esterases by substrate-imprinted docking

    Directory of Open Access Journals (Sweden)

    Tyagi Sadhna

    2009-06-01

    Full Text Available Abstract Background Previously, ways to adapt docking programs that were developed for modelling inhibitor-receptor interaction have been explored. Two main issues were discussed. First, when trying to model catalysis a reaction intermediate of the substrate is expected to provide more valid information than the ground state of the substrate. Second, the incorporation of protein flexibility is essential for reliable predictions. Results Here we present a predictive and robust method to model substrate specificity and enantioselectivity of lipases and esterases that uses reaction intermediates and incorporates protein flexibility. Substrate-imprinted docking starts with covalent docking of reaction intermediates, followed by geometry optimisation of the resulting enzyme-substrate complex. After a second round of docking the same substrate into the geometry-optimised structures, productive poses are identified by geometric filter criteria and ranked by their docking scores. Substrate-imprinted docking was applied in order to model (i enantioselectivity of Candida antarctica lipase B and a W104A mutant, (ii enantioselectivity and substrate specificity of Candida rugosa lipase and Burkholderia cepacia lipase, and (iii substrate specificity of an acetyl- and a butyrylcholine esterase toward the substrates acetyl- and butyrylcholine. Conclusion The experimentally observed differences in selectivity and specificity of the enzymes were reproduced with an accuracy of 81%. The method was robust toward small differences in initial structures (different crystallisation conditions or a co-crystallised ligand, although large displacements of catalytic residues often resulted in substrate poses that did not pass the geometric filter criteria.

  15. Characterisation of esterases as potential biomarkers of pesticide exposure in the lugworm Arenicola marina (Annelida: Polychaeta)

    Energy Technology Data Exchange (ETDEWEB)

    Hannam, Marie L. [Ecotoxicology and Stress Biology Research Centre, School of Biological Sciences, University of Plymouth, Drake Circus, Plymouth, Devon, PL4 8AA (United Kingdom)], E-mail: marie.hannam@plymouth.ac.uk; Hagger, Josephine A.; Jones, Malcolm B.; Galloway, Tamara S. [Ecotoxicology and Stress Biology Research Centre, School of Biological Sciences, University of Plymouth, Drake Circus, Plymouth, Devon, PL4 8AA (United Kingdom)

    2008-03-15

    Here, we identify and characterise cholinesterase (ChE) and carboxylesterase (CbE) activities in the body tissues of the sediment dwelling worm Arenicola marina. Exposure to the organophosphorus pesticide azamethiphos yielded an in vitro IC{sub 50} of 5 {mu}g l{sup -1} for propionylcholinesterase (PChE). PChE was significantly inhibited in vivo after a 10 day exposure to 100 {mu}g l{sup -1} azamethiphos, equivalent to the recommended aquatic application rate (ANOVA; F = 2.75, P = 0.033). To determine sensitivity to environmental conditions, A. marina were exposed for 10 days to field collected sediments. PChE activity was significantly lower in worms exposed to sediments from an estuary classified to be at high risk from point source pollution by the UK Environment Agency (ANOVA; F = 15.33, P < 0.001). Whilst causality cannot be directly attributed from these latter exposures, they provide an important illustration of the potential utility of esterase activity as a biomarker of environmental quality in this ecologically relevant sentinel species. - This paper provides a preliminary characterisation of esterase enzyme activities in the tissues and body fluids of the sediment dwelling worm Arenicola marina and explores their potential use as biomarkers of organophosphorus pesticide exposure in the marine environment.

  16. The MIEL1 E3 Ubiquitin Ligase Negatively Regulates Cuticular Wax Biosynthesis in Arabidopsis Stems.

    Science.gov (United States)

    Lee, Hong Gil; Kim, Juyoung; Suh, Mi Chung; Seo, Pil Joon

    2017-07-01

    Cuticular wax is an important hydrophobic layer that covers the plant aerial surface. Cuticular wax biosynthesis is shaped by multiple layers of regulation. In particular, a pair of R2R3-type MYB transcription factors, MYB96 and MYB30, are known to be the main participants in cuticular wax accumulation. Here, we report that the MYB30-INTERACTING E3 LIGASE 1 (MIEL1) E3 ubiquitin ligase controls the protein stability of the two MYB transcription factors and thereby wax biosynthesis in Arabidopsis. MIEL1-deficient miel1 mutants exhibit increased wax accumulation in stems, with up-regulation of wax biosynthetic genes targeted by MYB96 and MYB30. Genetic analysis reveals that wax accumulation of the miel1 mutant is compromised by myb96 or myb30 mutation, but MYB96 is mainly epistatic to MIEL1, playing a predominant role in cuticular wax deposition. These observations indicate that the MIEL1-MYB96 module is important for balanced cuticular wax biosynthesis in developing inflorescence stems. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  17. Climate Change Communicators: The C3E3 Project

    Science.gov (United States)

    Sharif, H. O.; Joseph, J.

    2013-12-01

    The University of Texas at San Antonio (UTSA), San Antonio College (SAC), and the University of North Dakota (UND) have partnered with NASA to provide underrepresented undergraduates from UTSA, SAC, and other community colleges climate-related research and education experiences through the Climate Change Communication: Engineer, Environmental science, and Education (C3E3) project. The program aims to develop a robust response to climate change by providing K-16 climate change education; enhance the effectiveness of K-16 education particularly in engineering and other STEM disciplines by use of new instructional technologies; increase the enrollment in engineering programs and the number of engineering degrees awarded by showing engineering's usefulness in relation to the much-discussed contemporary issue of climate change; increase persistence in STEM degrees by providing student research opportunities; and increase the ethnic diversity of those receiving engineering degrees and help ensure an ethnically diverse response to climate change. Students participated in the second summer internship funded by the project. More than 60 students participated in guided research experiences aligned with NASA Science Plan objectives for climate and Earth system science and the educational objectives of the three institutions. The students went through training in modern media technology (webcasts), and in using this technology to communicate the information on climate change to others, especially high school students, culminating in production of webcasts on investigating the aspects of climate change using NASA data. Content developed is leveraged by NASA Earth observation data and NASA Earth system models and tools. Several departments are involved in the educational program.

  18. Functional Characterization of the Apple RING E3 Ligase MdMIEL1 in Transgenic Arabidopsis

    Directory of Open Access Journals (Sweden)

    Jianping AN

    2017-03-01

    Full Text Available E3 ubiquitin ligases are involved in various physiological processes, and they play pivotal roles in growth and development. In this study, we identified a previously unknown gene in the apple fruit (Malus × domestica and named it MdMIEL1. The MdMIEL1 gene encoded a protein that contained a zinc-finger domain at its N-terminus and a RING-finger motif at its C-terminus. To investigate MdMIEL1 functions, we generated transgenic Arabidopsis lines expressing the MdMIEL1 gene under the control of the Cauliflower mosaic virus 35S promoter. Interestingly, ectopic expression of MdMIEL1 in Arabidopsis produced multiple phenotypes, including early germination, early flowering and a lateral root number increase relative to wild-type plants. Further analysis indicated that MdMIEL1 regulated lateral root initiation by increasing auxin accumulation in the roots. In a word, these results suggest that, MdMIEL1 as a novel RING-finger ubiquitin ligase influences plant growth and development, and highlight that MdMIEL1 regulates lateral root growth.

  19. TRIM E3 ligases interfere with early and late stages of the retroviral life cycle.

    Directory of Open Access Journals (Sweden)

    Pradeep D Uchil

    2008-02-01

    Full Text Available Members of the TRIpartite interaction Motif (TRIM family of E3 ligases have been shown to exhibit antiviral activities. Here we report a near comprehensive screen for antiretroviral activities of 55 TRIM proteins (36 human, 19 mouse. We identified approximately 20 TRIM proteins that, when transiently expressed in HEK293 cells, affect the entry or release of human immunodeficiency virus 1 (HIV, murine leukemia virus (MLV, or avian leukosis virus (ALV. While TRIM11 and 31 inhibited HIV entry, TRIM11 enhanced N-MLV entry by interfering with Ref1 restriction. Strikingly, many TRIM proteins affected late stages of the viral life cycle. Gene silencing of endogenously expressed TRIM 25, 31, and 62 inhibited viral release indicating that they play an important role at late stages of the viral life cycle. In contrast, downregulation of TRIM11 and 15 enhanced virus release suggesting that these proteins contribute to the endogenous restriction of retroviruses in cells.

  20. Mapping of repair genes

    International Nuclear Information System (INIS)

    Hori, Tadaaki

    1985-01-01

    Chromosome mapping of repair genes involved in U.V. sensitivity is reported. Twenty-three of 25 hybrid cells were resistant to U.V. light. Survival curves of 2 U.V.-resistant cell strains, which possessed mouse chromosomes and human chromosome No.7 - 16, were similar to those of wild strain (L5178Y). On the other hand, survival curves of U.V.-sensitive hybrid cells was analogous to those of Q31. There was a definitive difference in the frequency of inducible chromosome aberrations between U.V. resistant and sensitive mouse-human hybrid cells. U.V.-resistant cell strains possessed the ability of excision repair. Analysis of karyotype in hybrid cells showed that the difference in U.V. sensitivity is dependent upon whether or not human chromosome No.13 is present. Synteny test on esterase D-determining locus confirmed that there is an agreement between the presence of chromosome No.13 and the presence of human esterase D activity. These results led to a conclusion that human genes which compensate recessive character of U.V.-sensitive mutant strain, Q31, with mouse-human hybrid cells are located on the locus of chromosome No.13. (Namekawa, K.)

  1. In-gel detection of esterase-like albumin activity: Characterization of esterase-free sera albumin and its putative role as non-invasive biomarker of hepatic fibrosis

    Directory of Open Access Journals (Sweden)

    Areeba Ahmad

    2017-07-01

    Full Text Available Albumin is a globular and un-glycosylated multifunctional plasma protein and thus correlated with several human diseases. Owing to esterase contamination, albumin levels are usually misleading. In this study, we propose methodical accuracy for albumin estimation taking healthy and fibrotic rats. Liver fibrosis in rats was generated by N′-Nitrosodimethylamine (NDMA (10 mg/kg body weight within three weeks followed by its confirmation through H&E and immunohistochemical staining for α-SMA expression. Animal sera were screened by native polyacrylamide gel electrophoresis (native-PAGE (7.5%. In-gel esterase-like albumin activity was detected using α- and β-naphthyl acetate (5.58 × 10−3 mM; pH 7.5 as substrate. Sera albumin was purified from unstained PA gel-slices through electroelution. Subsequent to conformation of albumin purity by its molecular weight determination using SDS–PAGE (10% and peptide mass fingerprinting by MALDI-TOF-MS, samples were treated with different concentrations of urea. Urea-treated albumins were screened for esterase activity, conformational change and, albumin levels by immunoblotting. Our results demonstrate that esterase-like albumin activity in rat sera albumin is located in domain-III. The esterase-like activity remains detectable up to 4 M urea, which diminishes with increasing urea concentrations. Further, immunoblotting of urea-treated albumin samples displays a significant decline in purified protein bands, indicating hypoalbuminemia during hepatic fibrosis in rats. In conclusion, the present approach of albumin separation and estimation is of potential interest and may be recommended for diagnostic purposes.

  2. ISOLATION OF JUVENILE HORMONES ESTERASE AND ITS PARTIAL CDNA CLONE FROM THE BEETLE, TENEBRIO MOLITOR. (R825433)

    Science.gov (United States)

    Juvenile hormone esterase (JHE) plays an essential role in insect development. It is partially responsible for the clearance of juvenile hormone (JH) which regulates various aspects of insect development and reproduction. Because of its role in regulating JH titer, this enzyme...

  3. Effect of Temperature and High Pressure on the Activity and Mode of Action of Fungal Pectin Methyl Esterase

    NARCIS (Netherlands)

    Duvetter, T.; Fraeye, I.; Sila, D.N.; Verlent, I.; Smout, C.; Clynen, E.; Schoofs, L.; Schols, H.A.; Hendrickx, M.; Loey, van A.

    2006-01-01

    Pectin was de-esterified with purified recombinant Aspergillus aculeatus pectin methyl esterase (PME) during isothermal-isobaric treatments. By measuring the release of methanol as a function of treatment time, the rate of enzymatic pectin conversion was determined. Elevated temperature and pressure

  4. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm.

    Science.gov (United States)

    Harholt, Jesper; Bach, Inga C; Lind-Bouquin, Solveig; Nunan, Kylie J; Madrid, Susan M; Brinch-Pedersen, Henrik; Holm, Preben B; Scheller, Henrik V

    2010-04-01

    Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm-specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal (Lys-Asp-Glu-Leu) KDEL were used. Transgenic plants were recovered in all four cases but no qualitative differences could be observed whether KDEL was added or not. Endo-xylanase activity in transgenic grains was increased between two and threefold relative to wild type. The grains were shrivelled and had a 25%-33% decrease in mass. Extensive analysis of the cell walls showed a 10%-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water-extractable arabinoxylan, and a shift in the MW of the water-extractable arabinoxylan from being mainly larger than 85 kD to being between 2 and 85 kD. Ferulic acid esterase-expressing grains were also shrivelled, and the seed weight was decreased by 20%-50%. No ferulic acid esterase activity could be detected in wild-type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15%-40% increase in water-unextractable arabinoxylan and a decrease in monomeric ferulic acid between 13% and 34%. In all the plants, the observed changes are consistent with a plant response that serves to minimize the effect of the heterologously expressed enzymes by increasing arabinoxylan biosynthesis and cross-linking.

  5. Feruloyl esterases as a tool for the release of phenolic compounds from agro-industrial by-products

    NARCIS (Netherlands)

    Benoit, Isabelle; Navarro, David; Marnet, Nathalie; Rakotomanomana, Nnjara; Lesage-Meessen, Laurence; Sigoillot, Jean-Claude; Asther, Marcel; Asther, Michèle

    2006-01-01

    Agro-industrial by-products are a potential source of added-value phenolic acids with promising applications in the food and pharmaceutical industries. Here two purified feruloyl esterases from Aspergillus niger, FAEA and FAEB were tested for their ability to release phenolic acids such as caffeic

  6. Spatial distribution and esterase activity in populations of Aedes (Stegomyia aegypti (Linnaeus (Diptera: Culicidae resistant to temephos

    Directory of Open Access Journals (Sweden)

    Wanessa Porto Tito Gambarra

    2013-04-01

    Full Text Available INTRODUCTION: The need for studies that describe the resistance patterns in populations of Aedes aegypti (Linnaeus in function of their region of origin justified this research, which aimed to characterize the resistance to temephos and to obtain information on esterase activity in populations of Aedes aegypti collected in municipalities of the State of Paraíba. METHODS: Resistance to temephos was evaluated and characterized from the diagnostic dose of 0.352mg i.a./L and multiple concentrations that caused mortalities between 5% and 99%. Electrophoresis of isoenzymes was used to verify the patterns of esterase activity among populations of the vector. RESULTS: All populations of Aedes aegypti were resistant to temephos, presenting a resistance rate (RR greater than 20. The greatest lethal dose 50% of the sample (CL50 was found for the municipality of Lagoa Seca, approximately forty-one times the value of CL50 for the Rockefeller population. The populations characterized as resistant showed two to six regions of α and β-esterase, called EST-1 to EST-6, while the susceptible population was only seen in one region of activity. CONCLUSIONS: Aedes aegypti is widely distributed and shows a high degree of resistance to temephos in all municipalities studied. In all cases, esterases are involved in the metabolism and, consequently, in the resistance to temephos.

  7. Pig Liver Esterase (PLE) as Biocatalyst in Organic Synthesis: From Nature to Cloning and to Practical Applications

    NARCIS (Netherlands)

    Dominguez de Maria, Pablo; Garcia-Burgos, Carlos A.; Bargeman, Gerrald; van Gemert, Robert W.

    2007-01-01

    Pig liver esterase (PLE, EC 3.1.1.1) has been employed extensively for research purposes during the last three decades, especially in kinetic resolutions, in desymmetrizations of prochiral substrates, and in the synthesis of nucleosides. Its practical use, however, has been traditionally hampered

  8. Functional characterisation of a metagenome derived family VIII esterase with a deacetylation activity on ß-lactam antibiotics

    CSIR Research Space (South Africa)

    Mokoena, N

    2013-08-01

    Full Text Available characterisation of a novel esterase (Est22) derived from an acidic Leachate environment. The enzyme is 423 amino acids in length and contained 22aa signal peptide. Analysis of the Est22 primary structure revealed the presence of N-terminus S-x-x-K sequence, which...

  9. Crystallization and preliminary crystallographic analysis of an esterase with a novel domain from the hyperthermophile Thermotoga maritima

    NARCIS (Netherlands)

    Sun, Lei; Levisson, Mark; Hendriks, Sjon; Akveld, Twan; Kengen, Serve W. M.; Dijkstra, Bauke W.; van der Oost, John

    A predicted esterase ( EstA) with an unusual new domain from the hyperthermophilic bacterium Thermotoga maritima has been cloned and overexpressed in Escherichia coli. The purified protein was crystallized by the hanging-drop vapour-diffusion technique in the presence of lithium sulfate and

  10. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm

    DEFF Research Database (Denmark)

    Harholt, Jesper; Bach, Inga Christensen; Lind Bouquin, Solveig

    2010-01-01

    Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm-specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal (Lys-Asp-Glu-Leu) KDEL were used....... Extensive analysis of the cell walls showed a 10%-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water-extractable arabinoxylan, and a shift in the MW of the water-extractable arabinoxylan from being mainly larger than 85 kD to being between 2 and 85 kD. Ferulic acid esterase......-expressing grains were also shrivelled, and the seed weight was decreased by 20%-50%. No ferulic acid esterase activity could be detected in wild-type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15%-40% increase in water-unextractable arabinoxylan...

  11. Combining substrate specificity analysis with support vector classifiers reveals feruloyl esterase as a phylogenetically informative protein group

    DEFF Research Database (Denmark)

    Olivares Hernandez, Roberto; Sunner, Hampus; Frisvad, Jens Christian

    2010-01-01

    Background Our understanding of how fungi evolved to develop a variety of ecological niches, is limited but of fundamental biological importance. Specifically, the evolution of enzymes affects how well species can adapt to new environmental conditions. Feruloyl esterases (FAEs) are enzymes able...

  12. Evaluation of the nitrite and leukocyte esterase activity tests for the diagnosis of acute symptomatic urinary tract infection in men.

    NARCIS (Netherlands)

    Koeijers, J.J.; Kessels, A.G.H.; Nys, S.; Bartelds, A.; Donker, G.; Stobberingh, E.; Verbon, A.

    2007-01-01

    For 422 male patients with symptoms indicative of a urinary tract infection, nitrite and leukocyte esterase activity dipstick test results were compared with results of culture of urine samples. The positive predictive value of a positive nitrite test result was 96%. Addition of results of the

  13. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm

    Energy Technology Data Exchange (ETDEWEB)

    Harholt, Jesper; Bach, Inga C; Lind-Bouquin, Solveig; Nunan, Kylie J.; Madrid, Susan M.; Brinch-Pedersen, Henrik; Holm, Preben B.; Scheller, Henrik V.

    2009-12-08

    Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal KDEL were used. Transgenic plants were recovered in all four cases but no qualitative differences could be observed whether KDEL was added or not. Endo-xylanase activity in transgenic grains was increased between two and three fold relative to wild type. The grains were shriveled and had a 25-33% decrease in mass. Extensive analysis of the cell walls showed a 10-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water extractable arabinoxylan, and a shift in the MW of the water extractable arabinoxylan from being mainly larger than 85 kD to being between 2 kD and 85 kD. Ferulic acid esterase expressing grains were also shriveled and the seed weight was decreased by 20-50%. No ferulic acid esterase activity could be detected in wild type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15-40% increase in water unextractable arabinoxylan and a decrease in monomeric ferulic acid between 13 and 34%. In all the plants the observed changes are consistent with a plant response that serves to minimize the effect of the heterologously expressed enzymes by increasing arabinoxylan biosynthesis and cross-linking.

  14. Evaluating the efficacy of subcutaneous C1-esterase inhibitor administration for use in rat models of inflammatory diseases

    NARCIS (Netherlands)

    Emmens, Reindert W.; Naaijkens, Benno A.; Roem, Dorina; Kramer, Klaas; Wouters, Diana; Zeerleder, Sacha; van Ham, Marieke S.; Niessen, Hans W.; Krijnen, Paul A.

    2014-01-01

    Context: C1-esterase inhibitor (C1-inh) therapy is currently administered to patients with C1-inh deficiency through intravenous injections. The possibility of subcutaneous administration is currently being explored since this would alleviate need for hospitalization and increase mobility and

  15. Functional characterization of salt-tolerant microbial esterase WDEst17 and its use in the generation of optically pure ethyl (R)-3-hydroxybutyrate.

    Science.gov (United States)

    Wang, Yilong; Xu, Yongkai; Zhang, Yun; Sun, Aijun; Hu, Yunfeng

    2018-06-01

    The two enantiomers of ethyl 3-hydroxybutyrate are important intermediates for the synthesis of a great variety of valuable chiral drugs. The preparation of chiral drug intermediates through kinetic resolution reactions catalyzed by esterases/lipases has been demonstrated to be an efficient and environmentally friendly method. We previously functionally characterized microbial esterase PHE21 and used PHE21 as a biocatalyst to generate optically pure ethyl (S)-3-hydroxybutyrate. Herein, we also functionally characterized one novel salt-tolerant microbial esterase WDEst17 from the genome of Dactylosporangium aurantiacum subsp. Hamdenensis NRRL 18085. Esterase WDEst17 was further developed as an efficient biocatalyst to generate (R)-3-hydroxybutyrate, an important chiral drug intermediate, with the enantiomeric excess being 99% and the conversion rate being 65.05%, respectively, after process optimization. Notably, the enantio-selectivity of esterase WDEst17 was opposite than that of esterase PHE21. The identification of esterases WDEst17 and PHE21 through genome mining of microorganisms provides useful biocatalysts for the preparation of valuable chiral drug intermediates. © 2018 Wiley Periodicals, Inc.

  16. Esterases activity in the axolotl Ambystoma mexicanum exposed to chlorpyrifos and its implication to motor activity.

    Science.gov (United States)

    Robles-Mendoza, Cecilia; Zúñiga-Lagunes, Sebastian R; Ponce de León-Hill, Claudia A; Hernández-Soto, Jesús; Vanegas-Pérez, Cecilia

    2011-10-01

    The axolotl Ambystoma mexicanum is a neotenic salamander considered a good biological model due to its ability to regenerate limbs, tail, brain and heart cells. Nevertheless, severe reduction of A. mexicanum wild populations in the lacustrine area of Xochimilco, the natural habitat of the axolotl, could be related to several environmental pressures as the presence of organophosphate pesticides (OPPs), intensively applied in agricultural activities in Xochimilco. Thus the aim of this study was to evaluate the effect of environmentally realistic chlorpyrifos (CPF) concentrations, a OPP commonly used in this zone, on esterases activity (acetylcholinesterase and carboxylesterase) and bioconcentration of CPF and to relate them with the motor activity of A. mexicanum juveniles. Axolotls were exposed 48 h to 0.05 and 0.1mg CPF/L, and the responses were evaluated at the end of the CPF exposure. Results suggest that CPF is bioconcentrated into axolotls and that the CPF internal concentrations are related with the observed inhibition activity of AChE (>50%) and CbE (≈ 50%). CPF concentration responsible of the inhibition of the 50% of AChE activity (IC50) was estimated in 0.04 mg CPF/L; however IC50 for CbE activity was not possible to calculate since inhibition levels were lower than 50%, results that suggest a higher resistance of CbE enzymatic activity to CPF. However, motor activity was a more sensitive endpoint to CPF poisoning since time that axolotls spent active and walking, frequency and speed of swimming, frequency of prey attack were reduced >90% of control groups. The motor activity alterations in the axolotl could be related with the registered esterases inhibition. Thus important alterations on axolotls were identified even at short time and low concentrations of CPF exposure. Also, it was possible to link biochemical responses as esterases activity with higher levels of biological organization as behavior. This study provides tools for the regulation of the

  17. Analysing deltamethrin susceptibility and pyrethroid esterase activity variations in sylvatic and domestic Triatoma infestans at the embryonic stage

    Directory of Open Access Journals (Sweden)

    Pablo Luis Santo-Orihuela

    2013-12-01

    Full Text Available The aim of the present work was to study the deltamethrin susceptibility of eggs from Triatoma infestans populations and the contribution of pyrethroid esterases to deltamethrin degradation. Insects were collected from sylvatic areas, including Veinte de Octubre and Kirus-Mayu (Bolivia and from domiciliary areas, including El Palmar (Bolivia and La Pista (Argentina. Deltamethrin susceptibility was determined by dose-response bioassays. Serial dilutions of deltamethrin (0.0005-1 mg/mL were topically applied to 12-day-old eggs. Samples from El Palmar had the highest lethal dose ratio (LDR value (44.90 compared to the susceptible reference strain (NFS, whereas the Veinte de Octubre samples had the lowest value (0.50. Pyrethroid esterases were evaluated using 7-coumaryl permethrate (7-CP on individually homogenised eggs from each population and from NFS. The El Palmar and La Pista samples contained 40.11 and 36.64 pmol/min/mg protein, respectively, and these values were statistically similar to NFS (34.92 pmol/min/mg protein and different from Kirus-Mayu and Veinte de Octubre (27.49 and 22.69 pmol/min/mg protein, respectively. The toxicological data indicate that the domestic populations were resistant to deltamethrin, but no statistical contribution of 7-CP esterases was observed. The sylvatic populations had similar LDR values to NFS, but lower 7-CP esterase activities. Moreover, this is the first study of the pyrethroid esterases on T. infestans eggs employing a specific substrate (7-CP.

  18. Effects of temperature, ultraviolet radiation and pectin methyl esterase on aerobic methane release from plant material.

    Science.gov (United States)

    Bruhn, D; Mikkelsen, T N; Obro, J; Willats, W G T; Ambus, P

    2009-11-01

    This study examines the effects of different irradiance types on aerobic methane (CH(4)) efflux rates from terrestrial plant material. Furthermore, the role of the enzyme pectin methyl esterase (PME) on CH(4) efflux potential was also examined. Different types of plant tissue and purified pectin were incubated in glass vials with different combinations of irradiation and/or temperature. Purified dry pectin was incubated in solution, and with or without PME. Before and after incubation, the concentration of CH(4) was measured with a gas chromatograph. Rates of CH(4) emission were found to depend exponentially on temperature and linearly on UV-B irradiance. UV-B had a greater stimulating effect than UV-A, while visible light had no effect on emission rates. PME was found to substantially reduce the potential for aerobic CH(4) emissions upon demethylation of pectin.

  19. Synthesis activity-based zymography for detection of lipases and esterases.

    Science.gov (United States)

    Kwon, Min-A; Kim, Hyun Suk; Hahm, Dae-Hyun; Song, Jae Kwang

    2011-04-01

    A new zymography method for lipases and esterases was developed on the basis of the esterification reaction between fatty acids and alcohols. The enzymes were separated by SDS-PAGE and native PAGE. The gel was washed and then incubated in an aqueous solution containing fatty acids (oleic acid 18:1 or caprylic acid 8:0) and dodecanol. Synthesis was visualized by in situ precipitation of water-insoluble and non-diffusible fatty acid esters, such as dodecyl oleate and dodecyl octanoate. The synthesis activity-based zymography was confirmed with different enzyme samples, including commercial lipase preparations, purified recombinant lipase and cutinase, and crude culture supernatants of lipolytic enzyme-producing soil bacteria.

  20. Reactivity of Acetylcholine Esterase in inner Ear Maculae of Fish after Development at Hypergravity

    Science.gov (United States)

    Feucht, I.; Hilbig, R.; Anken, R.

    It has been shown earlier that the growth of inner ear otoliths of larval fish is (among other environmental factors) guided by the gravity vector. This guidance most probably is effected by the efferent vestibular system in the brainstem, because a transection of the nervus vestibularis has been shown to effect a cessation of the supply of calcium to the otoliths. The efferent innervation of fish inner ear maculae uses the synaptic transmitter acetylcholine (ACh). Therefore, we were - in order to further assess the role of the efferent system for otolith growth - prompted to determine ACh esterase-reactivity in the sensory epithelium of the utricle and the saccule (as well as in a non-gravity relevant brain region for control) in larval cichlid fish (Oreochromis mossambicus), which had been maintained at hypergravity during their development. The respective data will be communicated at the meeting. Acknowledgement: This work was financially supported by the German Aerospace Center (DLR) (FKZ: 50 WB 9997).

  1. Exceptional thermal stability and organic solvent tolerance of an esterase expressed from a thermophilic host

    DEFF Research Database (Denmark)

    Mei, Yuxia; Peng, Nan; Zhao, Shumiao

    2012-01-01

    , giving SisEstA. Upon Escherichia coli expression, only the thioredoxin-tagged EstA recombinant protein was soluble. The fusion protein was then purified, and removing the protein tag yielded EcSisEstA. Both forms of the thermophilic EstA enzyme were characterized. We found that SisEstA formed dimer...... that of EcSisEstA at 90°C. This indicated that thermophilic enzymes yielded from homologous expression should be better biocatalysts than those obtained from mesophilic expression.......A protein expression system recently developed for the thermophilic crenarchaeon Sulfolobus islandicus was employed to produce recombinant protein for EstA, a thermophilic esterase encoded in the same organism. Large amounts of protein were readily obtained by an affinity protein purification...

  2. From Classical to High Throughput Screening Methods for Feruloyl Esterases: A Review.

    Science.gov (United States)

    Ramírez-Velasco, Lorena; Armendáriz-Ruiz, Mariana; Rodríguez-González, Jorge Alberto; Müller-Santos, Marcelo; Asaff-Torres, Ali; Mateos-Díaz, Juan Carlos

    2016-01-01

    Feruloyl esterases (FAEs) are a diverse group of hydrolases widely distributed in plants and microorganisms which catalyzes the cleavage and formation of ester bonds between plant cell wall polysaccharides and phenolic acids. FAEs have gained importance in biofuel, medicine and food industries due to their capability of acting on a large range of substrates for cleaving ester bonds and synthesizing highadded value molecules through esterification and transesterification reactions. During the past two decades extensive studies have been carried out on the production, characterization and classification of FAEs, however only a few reports of suitable High Throughput Screening assays for this kind of enzymes have been reported. This review is focused on a concise but complete revision of classical to High Throughput Screening methods for FAEs, highlighting its advantages and disadvantages, and finally suggesting future perspectives for this important research field.

  3. Effect of phenobarbital on inducing insecticide tolerance and esterase changes in Aedes aegypti (Diptera: Culicidae

    Directory of Open Access Journals (Sweden)

    Rita de Cássia Sousa-Polezzi

    2004-01-01

    Full Text Available The effect of phenobarbital (PB on the induction of tolerance to the organophosphorous insecticide temephos (TE was investigated in Aedes aegypti L4 larvae submitted to two different PB-treatments:(1 continuous treatment from the egg to the larval L4 stage and (2 discontinuous treatment in which L4 larvae were exposed for 30 h. Mosquitoes from two Brazilian cities were studied: São José do Rio Preto (SJ in São Paulo State and Goiânia (GO in Goiás State. According to criterions established by World Health Organization (WHO mosquitoes from SJ are organophosphate-susceptible while mosquitoes from GO are organophosphate-resistant. For both SJ and GO larvae the two different PB-treatments resulted in significantly increased tolerance (measured by reduced mortality to 0.01mg/L TE while for larvae exposed to 0.02 mg/L TE only continuous PB-treatment resulted in significantly increased TE-tolerance. The reduction of mortality rate was greater in SJ larvae than in GO larvae, confirming data from other organisms indicating that the effect of PB is more pronounced in susceptible strains. To test if oxidase enzymes were involved in PB-induced tolerance we treated PB-pretreated SJ and GO larvae with the oxidase inhibitor piperonyl butoxide (PBO before exposure to TE and observed increased (rather than decreased tolerance, suggesting that oxidases are not involved in the tolerance process and that PB and PBO can act in concert or synergistically. Esterase patterns of PB-pretreated larvae indicated that the cholinesterases EST-13 and EST-14 are involved in the PB-induced TE- tolerance, reinforcing a previous study carried out in our laboratory which suggested that increased esterase synthesis is the mechanism responsible for the development of insecticide resistance in Aedes aegypti.

  4. Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading (TED)

    Science.gov (United States)

    Samtleben, Samira; Jaepel, Juliane; Fecher, Caroline; Andreska, Thomas; Rehberg, Markus; Blum, Robert

    2013-01-01

    Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca2+ indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca2+ indicator dyes in the ER lumen of different cell types. To date, TED was used in cell lines, glial cells, and neurons in vitro. TED bases on efficient, recombinant targeting of a high carboxylesterase activity to the ER lumen using vector-constructs that express Carboxylesterases (CES). The latest TED vectors contain a core element of CES2 fused to a red fluorescent protein, thus enabling simultaneous two-color imaging. The dynamics of free calcium in the ER are imaged in one color, while the corresponding ER structure appears in red. At the beginning of the procedure, cells are transduced with a lentivirus. Subsequently, the infected cells are seeded on coverslips to finally enable live cell imaging. Then, living cells are incubated with the acetoxymethyl ester (AM-ester) form of low-affinity Ca2+ indicators, for instance Fluo5N-AM, Mag-Fluo4-AM, or Mag-Fura2-AM. The esterase activity in the ER cleaves off hydrophobic side chains from the AM form of the Ca2+ indicator and a hydrophilic fluorescent dye/Ca2+ complex is formed and trapped in the ER lumen. After dye loading, the cells are analyzed at an inverted confocal laser scanning microscope. Cells are continuously perfused with Ringer-like solutions and the ER calcium dynamics are directly visualized by time-lapse imaging. Calcium release from the ER is identified by a decrease in fluorescence intensity in regions of interest, whereas the refilling of the ER calcium store produces an increase in fluorescence intensity. Finally, the change in fluorescent intensity over time is determined by calculation of ΔF/F0. PMID:23685703

  5. Direct imaging of ER calcium with targeted-esterase induced dye loading (TED).

    Science.gov (United States)

    Samtleben, Samira; Jaepel, Juliane; Fecher, Caroline; Andreska, Thomas; Rehberg, Markus; Blum, Robert

    2013-05-07

    Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca(2+) indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca(2+) indicator dyes in the ER lumen of different cell types. To date, TED was used in cell lines, glial cells, and neurons in vitro. TED bases on efficient, recombinant targeting of a high carboxylesterase activity to the ER lumen using vector-constructs that express Carboxylesterases (CES). The latest TED vectors contain a core element of CES2 fused to a red fluorescent protein, thus enabling simultaneous two-color imaging. The dynamics of free calcium in the ER are imaged in one color, while the corresponding ER structure appears in red. At the beginning of the procedure, cells are transduced with a lentivirus. Subsequently, the infected cells are seeded on coverslips to finally enable live cell imaging. Then, living cells are incubated with the acetoxymethyl ester (AM-ester) form of low-affinity Ca(2+) indicators, for instance Fluo5N-AM, Mag-Fluo4-AM, or Mag-Fura2-AM. The esterase activity in the ER cleaves off hydrophobic side chains from the AM form of the Ca(2+) indicator and a hydrophilic fluorescent dye/Ca(2+) complex is formed and trapped in the ER lumen. After dye loading, the cells are analyzed at an inverted confocal laser scanning microscope. Cells are continuously perfused with Ringer-like solutions and the ER calcium dynamics are directly visualized by time-lapse imaging. Calcium release from the ER is identified by a decrease in fluorescence intensity in regions of interest, whereas the refilling of the ER calcium store produces an increase in fluorescence intensity. Finally, the change in fluorescent intensity over time is determined by calculation of ΔF/F0.

  6. Novel organic solvent-tolerant esterase isolated by metagenomics: insights into the lipase/esterase classification Nueva esterasa tolerante a los solventes orgánicos aislada por metagenómica: ideas sobre la clasificación de las esterasas/lipasas

    Directory of Open Access Journals (Sweden)

    Renaud Berlemont

    2013-03-01

    Full Text Available In order to isolate novel organic solvent-tolerant (OST lipases, a metagenomic library was built using DNA derived from a temperate forest soil sample. A two-step activity-based screening allowed the isolation of a lipolytic clone active in the presence of organic solvents. Sequencing of the plasmid pRBest recovered from the positive clone revealed the presence of a putative lipase/esterase encoding gene. The deduced amino acid sequence (RBest1 contains the conserved lipolytic enzyme signature and is related to the previously described OST lipase from Lysinibacillus sphaericus 205y, which is the sole studied prokaryotic enzyme belonging to the 4.4 a/ß hydrolase subgroup (abH04.04. Both in vivo and in vitro studies of the substrate specificity of RBest1, using triacylglycerols or nitrophenyl-esters, respectively, revealed that the enzyme is highly specific for butyrate (C4 compounds, behaving as an esterase rather than a lipase. The RBest1 esterase was purified and biochemically characterized. The optimal esterase activity was observed at pH 6.5 and at temperatures ranging from 38 to 45 °C. Enzymatic activity, determined by hydrolysis of p-nitrophenyl esters, was found to be affected by the presence of different miscible and non-miscible organic solvents, and salts. Noteworthy, RBest1 remains significantly active at high ionic strength. These findings suggest that RBest1 possesses the ability of OST enzymes to molecular adaptation in the presence of organic compounds and resistance of halophilic proteins.Con el fin de aislar nuevas variantes de lipasas tolerantes a solventes organicos (OST, se construyo una libreria metagenomica a partir de ADN obtenido de una muestra de suelo de bosque templado. A traves de un monitoreo en dos etapas, basado en la deteccion de actividades, se aislo un clon con actividad lipolitica en presencia de solventes organicos. La secuenciacion del plasmido pRBest recuperado del clon positivo revelo la presencia de un

  7. Genome-wide and functional annotation of human E3 ubiquitin ligases identifies MULAN, a mitochondrial E3 that regulates the organelle's dynamics and signaling.

    Directory of Open Access Journals (Sweden)

    Wei Li

    2008-01-01

    Full Text Available Specificity of protein ubiquitylation is conferred by E3 ubiquitin (Ub ligases. We have annotated approximately 617 putative E3s and substrate-recognition subunits of E3 complexes encoded in the human genome. The limited knowledge of the function of members of the large E3 superfamily prompted us to generate genome-wide E3 cDNA and RNAi expression libraries designed for functional screening. An imaging-based screen using these libraries to identify E3s that regulate mitochondrial dynamics uncovered MULAN/FLJ12875, a RING finger protein whose ectopic expression and knockdown both interfered with mitochondrial trafficking and morphology. We found that MULAN is a mitochondrial protein - two transmembrane domains mediate its localization to the organelle's outer membrane. MULAN is oriented such that its E3-active, C-terminal RING finger is exposed to the cytosol, where it has access to other components of the Ub system. Both an intact RING finger and the correct subcellular localization were required for regulation of mitochondrial dynamics, suggesting that MULAN's downstream effectors are proteins that are either integral to, or associated with, mitochondria and that become modified with Ub. Interestingly, MULAN had previously been identified as an activator of NF-kappaB, thus providing a link between mitochondrial dynamics and mitochondria-to-nucleus signaling. These findings suggest the existence of a new, Ub-mediated mechanism responsible for integration of mitochondria into the cellular environment.

  8. The activity of non-specific esterase in the thyroid epithelial cells of the guinea pig as influenced by various inhibitors and activators. A histochemical study

    DEFF Research Database (Denmark)

    Kirkeby, S

    1976-01-01

    The action of various inhibitors and activators upon esterase activity in the thyroid epithelial cells is demonstrated. The agents used were triorthocresylphosphate (TOCP), parachloromercuribenzoate (PCMB), Arsanillic acid, p-nitrophenyl dimethyl carbamate and bis p-nitrophenyl phosphate. TOCP wa...

  9. Lactobacillus fermentum CRL1446 Ameliorates Oxidative and Metabolic Parameters by Increasing Intestinal Feruloyl Esterase Activity and Modulating Microbiota in Caloric-Restricted Mice

    Directory of Open Access Journals (Sweden)

    Matias Russo

    2016-07-01

    Full Text Available The purpose of this study was to determine whether the administration of the feruloyl esterase (FE-producing strain Lactobacillus fermentum CRL1446 enhances metabolic and oxidative parameters in caloric-restricted (CR mice. Balb/c male mice were divided into ad libitum fed Group (ALF Group, CR diet Group (CR Group and CR diet plus L. fermentum Group (CR-Lf Group. CR diet was administered during 45 days and CRL1446 strain was given in the dose of 108 cells/mL/day/mouse. FE activity was determined in intestinal mucosa and content at Day 1, 20 and 45. Triglyceride, total cholesterol, glucose, thiobarbituric acid reactive substances (TBARS levels and glutathione reductase activity were determined in plasma. Gut microbiota was evaluated by high-throughput sequencing of 16S rRNA gene amplicons. At Day 45, total intestinal FE activity in CR-Lf Group was higher (p = 0.020 than in CR and ALF groups and an improvement in both metabolic (reductions in triglyceride (p = 0.0025, total cholesterol (p = 0.005 and glucose (p < 0.0001 levels and oxidative (decrease of TBARS levels and increase of plasmatic glutathione reductase activity (p = 0.006 parameters was observed, compared to ALF Group. CR diet increased abundance of Bacteroidetes and CRL1446 administration increased abundance of Bifidobacterium and Lactobacillus genus. L. fermentun CRL1446 exerted a bifidogenic effect under CR conditions.

  10. Heterologous expression, purification and characterization of three novel esterases secreted by the lignocellulolytic fungus Penicillium purpurogenum when grown on sugar beet pulp.

    Science.gov (United States)

    Oleas, Gabriela; Callegari, Eduardo; Sepúlveda, Romina; Eyzaguirre, Jaime

    2017-04-18

    The lignocellulolytic fungus, Penicillium purpurogenum, grows on a variety of natural carbon sources, among them sugar beet pulp. Culture supernatants of P. purpurogenum grown on sugar beet pulp were partially purified and the fractions obtained analyzed for esterase activity by zymograms. The bands with activity on methyl umbelliferyl acetate were subjected to mass spectrometry to identify peptides. The peptides obtained were probed against the proteins deduced from the genome sequence of P. purpurogenum. Eight putative esterases thus identified were chosen for future work. Their cDNAs were expressed in Pichia pastoris. The supernatants of the recombinant clones were assayed for esterase activity, and five of the proteins were active against one or more substrates: methyl umbelliferyl acetate, indoxyl acetate, methyl esterified pectin and fluorescein diacetate. Three of those enzymes were purified, further characterized and subjected to a BLAST search. Based on their amino acid sequence and properties, they were identified as follows: RAE1, pectin acetyl esterase (CAZy family CE 12); FAEA, feruloyl esterase (could not be assigned to a CAZy family) and EAN, acetyl esterase (former CAZy family CE 10). Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Isolation and characterization of a heavy metal-resistant, thermophilic esterase from a Red Sea Brine Pool

    KAUST Repository

    Mohamed, Yasmine M.; Ghazy, Mohamed A.; Sayed, Ahmed; Ouf, Amged; El-Dorry, Hamza; Siam, Rania

    2013-01-01

    The Red Sea Atlantis II brine pool is an extreme environment that displays multiple harsh conditions such as high temperature, high salinity and high concentrations of multiple, toxic heavy metals. The survival of microbes in such an environment by utilizing resistant enzymes makes them an excellent source of extremophilic enzymes. We constructed a fosmid metagenomic library using DNA isolated from the deepest and most secluded layer of this pool. We report the isolation and biochemical characterization of an unusual esterase: EstATII. EstATII is thermophilic (optimum temperature, 65 C), halotolerant (maintains its activity in up to 4.5â€...M NaCl) and maintains at least 60% of its activity in the presence of a wide spectrum of heavy metals. The combination of biochemical characteristics of the Red Sea Atlantis II brine pool esterase, i.e., halotolerance, thermophilicity and resistance to heavy metals, makes it a potentially useful biocatalyst.

  12. Isolation and characterization of a heavy metal-resistant, thermophilic esterase from a Red Sea Brine Pool

    KAUST Repository

    Mohamed, Yasmine M.

    2013-11-28

    The Red Sea Atlantis II brine pool is an extreme environment that displays multiple harsh conditions such as high temperature, high salinity and high concentrations of multiple, toxic heavy metals. The survival of microbes in such an environment by utilizing resistant enzymes makes them an excellent source of extremophilic enzymes. We constructed a fosmid metagenomic library using DNA isolated from the deepest and most secluded layer of this pool. We report the isolation and biochemical characterization of an unusual esterase: EstATII. EstATII is thermophilic (optimum temperature, 65 C), halotolerant (maintains its activity in up to 4.5â€...M NaCl) and maintains at least 60% of its activity in the presence of a wide spectrum of heavy metals. The combination of biochemical characteristics of the Red Sea Atlantis II brine pool esterase, i.e., halotolerance, thermophilicity and resistance to heavy metals, makes it a potentially useful biocatalyst.

  13. Whole-Cell Biocatalytic Synthesis of Cinnamyl Acetate with a Novel Esterase from the DNA Library of Acinetobacter hemolyticus.

    Science.gov (United States)

    Dong, Hao; Secundo, Francesco; Xue, Changhu; Mao, Xiangzhao

    2017-03-15

    Cinnamyl acetate has a wide application in the flavor and fragrance industry because of its sweet, balsamic, and floral odor. Up to now, lipases have been mainly used in enzyme-mediated synthesis of cinnamyl acetate, whereas esterases are used in only a few cases. Moreover, the use of purified enzymes is often a disadvantage, which leads to increases of the production costs. In this paper, a genomic DNA library of Acinetobacter hemolyticus was constructed, and a novel esterase (EstK1) was identified. After expression in Escherichia coli, the whole-cell catalyst of EstK1 displayed high transesterification activity to produce cinnamyl acetate in nonaqueous systems. Furthermore, under optimal conditions (vinyl acetate as acyl donor, isooctane as solvent, molar ratio 1:4, temperature 40 °C), the conversion ratio of cinnamyl alcohol could be up to 94.1% at 1 h, and it reached an even higher level (97.1%) at 2 h.

  14. Isolated angioedema of the bowel due to C1 esterase inhibitor deficiency: a case report and review of literature

    Directory of Open Access Journals (Sweden)

    Kothari Shivangi T

    2011-02-01

    Full Text Available Abstract Introduction We report a rare, classic case of isolated angioedema of the bowel due to C1-esterase inhibitor deficiency. It is a rare presentation and very few cases have been reported worldwide. Angioedema has been classified into three categories. Case presentation A 66-year-old Caucasian man presented with a ten-month history of episodic severe cramping abdominal pain, associated with loose stools. A colonoscopy performed during an acute attack revealed nonspecific colitis. Computed tomography of the abdomen performed at the same time showed a thickened small bowel and ascending colon with a moderate amount of free fluid in the abdomen. Levels of C4 ( Conclusion In addition to a detailed comprehensive medical history, laboratory data and imaging studies are required to confirm a diagnosis of angioedema due to C1 esterase inhibitor deficiency.

  15. Identification of a Marine Bacillus Strain C5 and Parathion-Methyl Degradation Characteristics of the Extracellular Esterase B1

    Directory of Open Access Journals (Sweden)

    Jianhua Hao

    2014-01-01

    Full Text Available A bacterial strain C5 that can produce new type of marine esterase was isolated and screened from marine sludge. According to 16S rRNA sequence analysis and physiological and biochemical experiments, the strain was identified as Bacillus subtilis. A single isozyme with a molecular weight of 86 kDa was observed by SDS-PAGE and native-PAGE. On this basis, the mechanism of esterase B1 secreted by strain C5 degrading parathion-methyl was explored, and the effects of temperature and pH on the degradation rate were investigated. From the results, p-nitrophenol was one of the degradation products of B1 degrading parathion-methyl, and the best degradation effect could be achieved at the temperature of 40°C and the neutral pH value.

  16. The putative E3 ubiquitin ligase ECERIFERUM9 regulates abscisic acid biosynthesis and response during seed germination and postgermination growth in arabidopsis

    KAUST Repository

    Zhao, Huayan; Zhang, Huoming; Cui, Peng; Ding, Feng; Wang, Guangchao; Li, Rongjun; Jenks, Matthew A.; Lü , Shiyou; Xiong, Liming

    2014-01-01

    The ECERIFERUM9 (CER9) gene encodes a putative E3 ubiquitin ligase that functions in cuticle biosynthesis and the maintenance of plant water status. Here, we found that CER9 is also involved in abscisic acid (ABA) signaling in seeds and young

  17. Est16, a New Esterase Isolated from a Metagenomic Library of a Microbial Consortium Specializing in Diesel Oil Degradation.

    Science.gov (United States)

    Pereira, Mariana Rangel; Mercaldi, Gustavo Fernando; Maester, Thaís Carvalho; Balan, Andrea; Lemos, Eliana Gertrudes de Macedo

    2015-01-01

    Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1) from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404). The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes.

  18. Effects of high hydrostatic pressure and temperature increase on Escherichia coli spp. and pectin methyl esterase inactivation in orange juice.

    Science.gov (United States)

    Torres, E F; González-M, G; Klotz, B; Rodrigo, D

    2016-03-01

    The aim of this study was to evaluate the effect of high hydrostatic pressure treatment combined with moderate processing temperatures (25 ℃-50 ℃) on the inactivation of Escherichia coli O157: H7 (ATCC 700728), E. coli K12 (ATCC 23716), and pectin methyl esterase in orange juice, using pressures of 250 to 500 MPa with times ranging between 1 and 30 min. Loss of viability of E. coli O157:H7 increased significantly as pressure and treatment time increased, achieving a 6.5 log cycle reduction at 400 MPa for 3 min at 25 ℃ of treatment. With regard to the inactivation of pectin methyl esterase, the greatest reduction obtained was 90.05 ± 0.01% at 50 ℃ and 500 MPa of pressure for 15 min; therefore, the pectin methyl esterase enzyme was highly resistant to the treatments by high hydrostatic pressure. The results obtained in this study showed a synergistic effect between the high pressure and moderate temperatures in inactivating E. coli cells. © The Author(s) 2016.

  19. Ferulic acid: an antioxidant found naturally in plant cell walls and feruloyl esterases involved in its release and their applications.

    Science.gov (United States)

    Mathew, Sindhu; Abraham, T Emilia

    2004-01-01

    Ferulic acid is the most abundant hydroxycinnamic acid in the plant world and maize bran with 3.1% (w/w) ferulic acid is one of the most promising sources of this antioxidant. The dehydrodimers of ferulic acid are important structural components in the plant cell wall and serve to enhance its rigidity and strength. Feruloyl esterases are a subclass of the carboxylic acid esterases that hydrolyze the ester bond between hydroxycinnamic acids and sugars present in plant cell walls and they have been isolated from a wide range of microorganisms, when grown on complex substrates such as cereal brans, sugar beet pulp, pectin and xylan. These enzymes perform a function similar to alkali in the deesterification of plant cell wall and differ in their specificities towards the methyl esters of cinnamic acids and ferulolylated oligosaccharides. They act synergistically with xylanases and pectinases and facilitate the access of hydrolases to the backbone of cell wall polymers. The applications of ferulic acid and feruloyl esterase enzymes are many and varied. Ferulic acid obtained from agricultural byproducts is a potential precursor for the production of natural vanillin, due to the lower production cost.

  20. A cold active (2R,3R)-(-)-di-O-benzoyl-tartrate hydrolyzing esterase from Rhodotorula mucilaginosa.

    Science.gov (United States)

    Zimmer, Christian; Platz, Tanja; Cadez, Neza; Giffhorn, Friedrich; Kohring, Gert-Wieland

    2006-11-01

    In a screening procedure a pink-colored yeast was isolated from enrichment cultures with (2R,3R)-(-)-di-O-benzoyl-tartrate (benzoyl-tartrate) as the sole carbon source. The organism saar1 was identified by morphological, physiological, and 18S ribosomal DNA/internal transcribed spacer analysis as Rhodotorula mucilaginosa, a basidiomycetous yeast. During growth the yeast hydrolyzed the dibenzoyl ester stoichiometrically to the monoester using the separated benzoate as the growth substrate, before the monoester was further cleaved into benzoate and tartrate, which were both metabolized. The corresponding benzoyl esterase was purified from the culture supernatant and characterized as a monomeric glycosylated 86-kDa protein with an optimum pH of 7.5 and an optimum temperature of 45 degrees C. At 0 degrees C the esterase still exhibited 20% of the corresponding activity at 30 degrees C, which correlates it to psychrophilic enzymes. The esterase could hydrolyze short chain p-nitrophenyl-alkyl esters and several benzoyl esters like benzoyl-methyl ester, ethylene-glycol-dibenzoyl ester, phenyl-benzoyl ester, cocaine, and 1,5-anhydro-D: -fructose-tribenzoyl ester. However feruloyl-ethyl ester was not hydrolyzed. The activity characteristics let the enzyme appear as a promising tool for synthesis of benzoylated compounds for pharmaceutical, cosmetic, or fine chemical applications, even at low temperatures.

  1. Est16, a New Esterase Isolated from a Metagenomic Library of a Microbial Consortium Specializing in Diesel Oil Degradation.

    Directory of Open Access Journals (Sweden)

    Mariana Rangel Pereira

    Full Text Available Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1 from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404. The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes.

  2. Solution Behavior and Activity of a Halophilic Esterase under High Salt Concentration

    Science.gov (United States)

    Rao, Lang; Zhao, Xiubo; Pan, Fang; Li, Yin; Xue, Yanfen; Ma, Yanhe; Lu, Jian R.

    2009-01-01

    Background Halophiles are extremophiles that thrive in environments with very high concentrations of salt. Although the salt reliance and physiology of these extremophiles have been widely investigated, the molecular working mechanisms of their enzymes under salty conditions have been little explored. Methodology/Principal Findings A halophilic esterolytic enzyme LipC derived from archeaon Haloarcula marismortui was overexpressed from Escherichia coli BL21. The purified enzyme showed a range of hydrolytic activity towards the substrates of p-nitrophenyl esters with different alkyl chains (n = 2−16), with the highest activity being observed for p-nitrophenyl acetate, consistent with the basic character of an esterase. The optimal esterase activities were found to be at pH 9.5 and [NaCl] = 3.4 M or [KCl] = 3.0 M and at around 45°C. Interestingly, the hydrolysis activity showed a clear reversibility against changes in salt concentration. At the ambient temperature of 22°C, enzyme systems working under the optimal salt concentrations were very stable against time. Increase in temperature increased the activity but reduced its stability. Circular dichroism (CD), dynamic light scattering (DLS) and small angle neutron scattering (SANS) were deployed to determine the physical states of LipC in solution. As the salt concentration increased, DLS revealed substantial increase in aggregate sizes, but CD measurements revealed the maximal retention of the α-helical structure at the salt concentration matching the optimal activity. These observations were supported by SANS analysis that revealed the highest proportion of unimers and dimers around the optimal salt concentration, although the coexistent larger aggregates showed a trend of increasing size with salt concentration, consistent with the DLS data. Conclusions/Significance The solution α-helical structure and activity relation also matched the highest proportion of enzyme unimers and dimers. Given that

  3. Shutdown of interferon signaling by a viral-hijacked E3 ubiquitin ligase

    Directory of Open Access Journals (Sweden)

    Kaitlin A. Davis

    2017-11-01

    Full Text Available Viruses manipulate cellular processes to create an environment favorable to replication. For most viruses, this includes subverting the expression of interferon (IFN, a signaling molecule that can stimulate production of a vast array of antiviral gene products. Rotavirus, a segmented double-stranded RNA virus that causes acute gastroenteritis in infants and young children, inhibits IFN expression through its nonstructural protein NSP1. This viral protein stifles IFN expression by inducing the degradation of host factors that are necessary for upregulating the activity of IFN genes. In the case of nearly all human and porcine rotavirus strains, NSP1 induces the ubiquitination-dependent proteasomal degradation of β-transducin repeat containing protein (β-TrCP, a host factor that plays an essential role in activating the IFN-transcription factor, NF-κB. Key to the process is the presence of a decoy sequence (degron at the C-terminus of NSP1 that causes β-TrCP to mistakenly bind NSP1 instead of its natural target, inhibitor-of-κB (IκB. In a recent report published by Davis et al [2017; mBio 8(4: e01213-17], we describe molecular requirements that govern NSP1 recognition of β-TrCP, including an essential degron phosphorylation event, and the step-wise incorporation of NSP1 into hijacked cullin-RING E3 ligases (CRLs that ubiquitinate and tag β-TrCP for degradation. Notably, although β-TrCP is chiefly recognized for its role as a master regulator of NF-κB signaling and IFN expression, β-TrCP also controls the stability of checkpoint proteins implicated in numerous other cellular pathways with antiviral activities, including autophagy and apoptosis. Thus, the impact of NSP1 on creating an intracellular environment favorable to virus replication may extend well beyond the IFN signaling pathway.

  4. Mutation in SUMO E3 ligase, SIZ1, disrupts the mature female gametophyte in Arabidopsis

    KAUST Repository

    Ling, Yu

    2012-01-09

    Female gametophyte is the multicellular haploid structure that can produce embryo and endosperm after fertilization, which has become an attractive model system for investigating molecular mechanisms in nuclei migration, cell specification, cell-to-cell communication and many other processes. Previous reports found that the small ubiquitin-like modifier (SUMO) E3 ligase, SIZ1, participated in many processes depending on particular target substrates and suppression of salicylic acid (SA) accumulation. Here, we report that SIZ1 mediates the reproductive process. SIZ1 showed enhanced expression in female organs, but was not detected in the anther or pollen. A defect in the siz1-2 maternal source resulted in reduced seed-set regardless of high SA concentration within the plant. Moreover, aniline blue staining and scanning electron microscopy revealed that funicular and micropylar pollen tube guidance was arrested in siz1-2 plants. Some of the embryo sacs of ovules in siz1-2 were also disrupted quickly after stage FG7. There was no significant affects of the siz1-2 mutation on expression of genes involved in female gametophyte development- or pollen tube guidance in ovaries. Together, our results suggest that SIZ1 sustains the stability and normal function of the mature female gametophyte which is necessary for pollen tube guidance. © 2012 Ling et al.

  5. Electronic and structural properties of B i2S e3:Cu

    Science.gov (United States)

    Sobczak, Kamil; Strak, Pawel; Kempisty, Pawel; Wolos, Agnieszka; Hruban, Andrzej; Materna, Andrzej; Borysiuk, Jolanta

    2018-04-01

    Electronic and structural properties of B i2S e3 and its extension to copper doped B i2S e3:Cu were studied using combined ab initio simulations and transmission electron microscopy based techniques, including electron energy loss spectroscopy, energy filtered transmission electron microscopy, and energy dispersive x-ray spectroscopy. The stability of the mixed phases was investigated for substitutional and intercalation changes of basic B i2S e3 structure. Four systems were compared: B i2S e3 , structures obtaining by Cu intercalation of the van der Waals gap, by substitution of Bi by Cu in quintuple layers, and C u2Se . The structures were identified and their electronic properties were obtained. Transmission electron microscopy measurements of B i2S e3 and the B i2S e3:Cu system identified the first structure as uniform and the second as composite, consisting of a nonuniform lower-Cu-content matrix and randomly distributed high-Cu-concentration precipitates. Critical comparison of the ab initio and experimental data identified the matrix as having a B i2S e3 dominant part with randomly distributed Cu-intercalated regions having 1Cu-B i2S e3 structure. The precipitates were determined to have 3Cu-B i2S e3 structure.

  6. NR2E3 mutations in enhanced S-cone sensitivity syndrome (ESCS), Goldmann-Favre syndrome (GFS), clumped pigmentary retinal degeneration (CPRD), and retinitis pigmentosa (RP).

    Science.gov (United States)

    Schorderet, Daniel F; Escher, Pascal

    2009-11-01

    NR2E3, also called photoreceptor-specific nuclear receptor (PNR), is a transcription factor of the nuclear hormone receptor superfamily whose expression is uniquely restricted to photoreceptors. There, its physiological activity is essential for proper rod and cone photoreceptor development and maintenance. Thirty-two different mutations in NR2E3 have been identified in either homozygous or compound heterozygous state in the recessively inherited enhanced S-cone sensitivity syndrome (ESCS), Goldmann-Favre syndrome (GFS), and clumped pigmentary retinal degeneration (CPRD). The clinical phenotype common to all these patients is night blindness, rudimental or absent rod function, and hyperfunction of the "blue" S-cones. A single p.G56R mutation is inherited in a dominant manner and causes retinitis pigmentosa (RP). We have established a new locus-specific database for NR2E3 (www.LOVD.nl/eye), containing all reported mutations, polymorphisms, and unclassified sequence variants, including novel ones. A high proportion of mutations are located in the evolutionarily-conserved DNA-binding domains (DBDs) and ligand-binding domains (LBDs) of NR2E3. Based on homology modeling of these NR2E3 domains, we propose a structural localization of mutated residues. The high variability of clinical phenotypes observed in patients affected by NR2E3-linked retinal degenerations may be caused by different disease mechanisms, including absence of DNA-binding, altered interactions with transcriptional coregulators, and differential activity of modifier genes.

  7. The role of low levels of juvenile hormone Esterase in the metamorphosis of Manduca sexta

    Directory of Open Access Journals (Sweden)

    M.H. Browder

    2001-10-01

    Full Text Available The activity of juvenile hormone esterase (JHE in feeding fifth instar larvae of Manduca sexta increases gradually with larval weight and rises to a peak after larvae pass the critical weight when juvenile hormone secretion ceases. Starvation of larvae of Manduca sexta (L. that had exceeded the critical weight inhibited peak levels of JHE, but did not delay entry into the wandering stage when larvae leave the plant in search of a pupation site. This suggests that peak levels of JHE may not be essential for the normal timing of metamorphosis. Starved larvae pupated normally, indicating the peak of JHE was not necessary for a morphologically normal pupation. Treatments of larvae with the selective JHE inhibitor O-ethyl-S-phenyl phosphoramidothiolate (EPPAT that began immediately after larvae achieved the critical weight (6.0 to 6.5 grams for our strain of Manduca delayed entry into the wandering stage. By contrast, EPPAT treatment of larvae at weights above 8.0g had no effect on the subsequent timing of the onset of wandering. Therefore, although the normal timing of the onset of wandering does not require peak levels of JHE, it requires low to moderate levels of JHE to be present until larvae reach a weight of about 8.0g.

  8. Subunit Stabilization and Polyethylene Glycolation of Cocaine Esterase Improves In Vivo Residence Time

    Energy Technology Data Exchange (ETDEWEB)

    Narasimhan, Diwahar; Collins, Gregory T.; Nance, Mark R.; Nichols, Joseph; Edwald, Elin; Chan, Jimmy; Ko, Mei-Chuan; Woods, James H.; Tesmer, John J.G.; Sunahara, Roger K. (Michigan)

    2012-03-15

    No small-molecule therapeutic is available to treat cocaine addiction, but enzyme-based therapy to accelerate cocaine hydrolysis in serum has gained momentum. Bacterial cocaine esterase (CocE) is the fastest known native enzyme that hydrolyzes cocaine. However, its lability at 37 C has limited its therapeutic potential. Cross-linking subunits through disulfide bridging is commonly used to stabilize multimeric enzymes. Herein we use structural methods to guide the introduction of two cysteine residues within dimer interface of CocE to facilitate intermolecular disulfide bond formation. The disulfide-crosslinked enzyme displays improved thermostability, particularly when combined with previously described mutations that enhance stability (T172R-G173Q). The newly modified enzyme yielded an extremely stable form of CocE (CCRQ-CocE) that retained greater than 90% of its activity after 41 days at 37 C, representing an improvement of more than 4700-fold over the wild-type enzyme. CCRQ-CocE could also be modified by polyethylene glycol (PEG) polymers, which improved its in vivo residence time from 24 to 72 h, as measured by a cocaine lethality assay, by self-administration in rodents, and by measurement of inhibition of cocaine-induced cardiovascular effects in rhesus monkeys. PEG-CCRQ elicited negligible immune response in rodents. Subunit stabilization and PEGylation has thus produced a potential protein therapeutic with markedly higher stability both in vitro and in vivo.

  9. Pectin methyl esterase inhibits intrusive and symplastic cell growth in developing wood cells of Populus.

    Science.gov (United States)

    Siedlecka, Anna; Wiklund, Susanne; Péronne, Marie-Amélie; Micheli, Fabienne; Lesniewska, Joanna; Sethson, Ingmar; Edlund, Ulf; Richard, Luc; Sundberg, Björn; Mellerowicz, Ewa J

    2008-02-01

    Wood cells, unlike most other cells in plants, grow by a unique combination of intrusive and symplastic growth. Fibers grow in diameter by diffuse symplastic growth, but they elongate solely by intrusive apical growth penetrating the pectin-rich middle lamella that cements neighboring cells together. In contrast, vessel elements grow in diameter by a combination of intrusive and symplastic growth. We demonstrate that an abundant pectin methyl esterase (PME; EC 3.1.1.11) from wood-forming tissues of hybrid aspen (Populus tremula x tremuloides) acts as a negative regulator of both symplastic and intrusive growth of developing wood cells. When PttPME1 expression was up- and down-regulated in transgenic aspen trees, the PME activity in wood-forming tissues was correspondingly altered. PME removes methyl ester groups from homogalacturonan (HG) and transgenic trees had modified HG methylesterification patterns, as demonstrated by two-dimensional nuclear magnetic resonance and immunostaining using PAM1 and LM7 antibodies. In situ distributions of PAM1 and LM7 epitopes revealed changes in pectin methylesterification in transgenic trees that were specifically localized in expanding wood cells. The results show that en block deesterification of HG by PttPME1 inhibits both symplastic growth and intrusive growth. PttPME1 is therefore involved in mechanisms determining fiber width and length in the wood of aspen trees.

  10. Pectin Methyl Esterase Activity Change in Intermediate Moisture Sun-Dried Figs after Storage

    Directory of Open Access Journals (Sweden)

    Dilek Demirbüker Kavak

    2015-12-01

    Full Text Available Intermediate moisture fruits can be obtained by rehydrating dried fruits. Intermediate moisture fruits are suitable for direct consumption compared to dry fruits and can be directly used in the production of various products such as bakery products, dairy products and candies. Aim of this study is to compare the pectin methyl esterase (PME activity of intermediate moisture figs which causes softening of the texture and to compare their microbial stability after 3 months storage period. For this purpose, dried figs were rehydrated in 30 and 80° C water until they reach 30% moisture content. Rehydrated samples were stored for 3 months at +4°C. Results showed that there was no statistically significant difference between the control samples and the samples rehydrated at 80°C according to the total viable counts. At the end of the storage period, results of residual PME activity in control samples was 24.1 μmol COOH min-1g-1, while it was found 17.4 μmol COOH min-1g-1 in samples rehydrated at 80°C. As a result rehydration conducted at 80°C provided 28% reduction in PME activity compared to the control samples rehydrated at 30°C, although it did not affect the microbial load significantly after storage.

  11. An increase in pectin methyl esterase activity accompanies dormancy breakage and germination of yellow cedar seeds.

    Science.gov (United States)

    Ren, C; Kermode, A R

    2000-09-01

    Pectin methyl esterase (PME) (EC 3.1.1.11) catalyzes the hydrolysis of methylester groups of cell wall pectins. We investigated the role of this enzyme in dormancy termination and germination of yellow cedar (Chamaecyparis nootkatensis [D. Don] Spach) seeds. PME activity was not detected in dormant seeds of yellow cedar but was induced and gradually increased during moist chilling; high activity coincided with dormancy breakage and germination. PME activity was positively correlated to the degree of dormancy breakage of yellow cedar seeds. The enzyme produced in different seed parts and in seeds at different times during moist chilling, germination, and early post-germinative growth consisted of two isoforms, both basic with isoelectric points of 8.7 and 8.9 and the same molecular mass of 62 kD. The pH optimum for the enzyme was between 7.4 and 8.4. In intact yellow cedar seeds, activities of the two basic isoforms of PME that were induced in embryos and in megagametophytes following dormancy breakage were significantly suppressed by abscisic acid. Gibberellic acid had a stimulatory effect on the activities of these isoforms in embryos and megagametophytes of intact seeds at the germinative stage. We hypothesize that PME plays a role in weakening of the megagametophyte, allowing radicle emergence and the completion of germination.

  12. High-pressure processing of apple juice: kinetics of pectin methyl esterase inactivation.

    Science.gov (United States)

    Riahi, Esmaeil; Ramaswamy, Hosahalli S

    2003-01-01

    High-pressure (HP) inactivation kinetics of pectin methyl esterase (PME) in apple juice were evaluated. Commercial PME was dispensed in clarified apple juice, sealed in dual peel sterilizable plastic bags, and subjected to different high-pressure processing conditions (200-400 MPa, 0-180 min). Residual enzyme activity was determined by a titration method estimating the rate of free carboxyl group released by the enzyme acting on pectin substrate at pH 7.5 (30 degrees C). The effects of pressure level and pressure holding time on enzyme inactivation were significant (p < 0.05). PME from the microbial source was found to be more resistant (p < 0.05) to pressure inactivation than PME from the orange peel. Almost a full decimal reduction in the activity of commercial PME was achieved by HP treatment at 400 MPa for 25 min. Inactivation kinetics were evaluated on the basis of a dual effect model involving a pressure pulse effect and a first-order rate model, and the pressure sensitivity of rate constants was modeled by using the z-value concept.

  13. Separating esterase targets of organophosphorus compounds in the brain by preparative chromatography.

    Science.gov (United States)

    Mangas, I; Vilanova, E; Benabent, M; Estévez, J

    2014-02-10

    Low level exposure to organophosphorus esters (OPs) may cause long-term neurological effects and affect specific cognition domains in experimental animals and humans. Action on known targets cannot explain most of these effects by. Soluble carboxylesterases (EC 3.1.1.1) of chicken brain have been kinetically discriminated using paraoxon, mipafox and phenylmethyl sulfonylfluoride as inhibitors and phenyl valerate as a substrate. Three different enzymatic components were discriminated and called Eα, Eβ and Eγ. In this work, a fractionation procedure with various steps was developed using protein native separation methods by preparative HPLC. Gel permeation chromatography followed by ion exchange chromatography allowed enriched fractions with different kinetic behaviors. The soluble chicken brain fraction was fractionated, while total esterase activity, proteins and enzymatic components Eα, Eβ and Eγ were monitored in each subfraction. After the analysis, 13 fractions were pooled and conserved. Preincubation of the soluble chicken brain fraction of with the organophosphorus mipafox gave rise to a major change in the ion exchange chromatography profile, but not in the molecular exchanged chromatography profile, which suggest that mipafox permanently modifies the ionic properties of numerous proteins. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  14. Glucuronoyl Esterase Screening and Characterization Assays Utilizing Commercially Available Benzyl Glucuronic Acid Ester

    Directory of Open Access Journals (Sweden)

    Hampus Sunner

    2015-09-01

    Full Text Available Research on glucuronoyl esterases (GEs has been hampered by the lack of enzyme assays based on easily obtainable substrates. While benzyl d-glucuronic acid ester (BnGlcA is a commercially available substrate that can be used for GE assays, several considerations regarding substrate instability, limited solubility and low apparent affinities should be made. In this work we discuss the factors that are important when using BnGlcA for assaying GE activity and show how these can be applied when designing BnGlcA-based GE assays for different applications: a thin-layer chromatography assay for qualitative activity detection, a coupled-enzyme spectrophotometric assay that can be used for high-throughput screening or general activity determinations and a HPLC-based detection method allowing kinetic determinations. The three-level experimental procedure not merely facilitates routine, fast and simple biochemical characterizations but it can also give rise to the discovery of different GEs through an extensive screening of heterologous Genomic and Metagenomic expression libraries.

  15. Construction of an Immobilized Thermophilic Esterase on Epoxy Support for Poly(ε-caprolactone Synthesis

    Directory of Open Access Journals (Sweden)

    Hui Ren

    2016-06-01

    Full Text Available Developing an efficient immobilized enzyme is of great significance for improving the operational stability of enzymes in poly(ε-caprolactone synthesis. In this paper, a thermophilic esterase AFEST from the archaeon Archaeoglobus fulgidus was successfully immobilized on the epoxy support Sepabeads EC-EP via covalent attachment, and the immobilized enzyme was then employed as a biocatalyst for poly(ε-caprolactone synthesis. The enzyme loading and recovered activity of immobilized enzyme was measured to be 72 mg/g and 10.4 U/mg using p-nitrophenyl caprylate as the substrate at 80 °C, respectively. Through the optimization of reaction conditions (enzyme concentration, temperature, reaction time and medium, poly(ε-caprolactone was obtained with 100% monomer conversion and low number-average molecular weight (Mn < 1300 g/mol. Further, the immobilized enzyme exhibited excellent reusability, with monomer conversion values exceeding 75% during 15 batch reactions. Finally, poly(ε-caprolactone was enzymatically synthesized with an isolated yield of 75% and Mn value of 3005 g/mol in a gram-scale reaction.

  16. Construction of an Immobilized Thermophilic Esterase on Epoxy Support for Poly(ε-caprolactone) Synthesis.

    Science.gov (United States)

    Ren, Hui; Xing, Zhen; Yang, Jiebing; Jiang, Wei; Zhang, Gang; Tang, Jun; Li, Quanshun

    2016-06-18

    Developing an efficient immobilized enzyme is of great significance for improving the operational stability of enzymes in poly(ε-caprolactone) synthesis. In this paper, a thermophilic esterase AFEST from the archaeon Archaeoglobus fulgidus was successfully immobilized on the epoxy support Sepabeads EC-EP via covalent attachment, and the immobilized enzyme was then employed as a biocatalyst for poly(ε-caprolactone) synthesis. The enzyme loading and recovered activity of immobilized enzyme was measured to be 72 mg/g and 10.4 U/mg using p-nitrophenyl caprylate as the substrate at 80 °C, respectively. Through the optimization of reaction conditions (enzyme concentration, temperature, reaction time and medium), poly(ε-caprolactone) was obtained with 100% monomer conversion and low number-average molecular weight (Mn enzyme exhibited excellent reusability, with monomer conversion values exceeding 75% during 15 batch reactions. Finally, poly(ε-caprolactone) was enzymatically synthesized with an isolated yield of 75% and Mn value of 3005 g/mol in a gram-scale reaction.

  17. Design, characterisation and application of alginate-based encapsulated pig liver esterase.

    Science.gov (United States)

    Pauly, Jan; Gröger, Harald; Patel, Anant V

    2018-06-05

    Encapsulation of hydrolases in biopolymer-based hydrogels often suffers from low activities and encapsulation efficiencies along with high leaching and unsatisfactory recycling properties. Exemplified for the encapsulation of pig liver esterase the coating of alginate and chitosan beads have been studied by creating various biopolymer hydrogel beads. Enzyme activity and encapsulation efficiency were notably enhanced by chitosan coating of alginate beads while leaching remained nearly unchanged. This was caused by the enzymatic reaction acidifying the matrix, which increased enzyme retention through enhanced electrostatic enzyme-alginate interaction but decreased activity through enzyme deactivation. A practical and ready-to-use method for visualising pH in beads during reaction by co-encapsulation of a conventional pH indicator was also found. Our method proves that pH control inside the beads can only be realised by buffering. The resulting beads provided a specific activity of 0.267 μmol ∙ min -1 ∙ mg -1 , effectiveness factor 0.88, encapsulation efficiency of 88%, 5% leaching and good recycling properties. This work will contribute towards better understanding and application of encapsulated hydrolases for enzymatic syntheses. Copyright © 2018 Elsevier B.V. All rights reserved.

  18. Acetylcholine esterase activity in mild cognitive impairment and Alzheimer's disease

    Energy Technology Data Exchange (ETDEWEB)

    Herholz, Karl [University of Manchester, Wolfson Molecular Imaging Centre, Clinical Neuroscience, Manchester (United Kingdom); University of Cologne, Cologne (Germany)

    2008-03-15

    Impairment of cholinergic neurotransmission is a well-established fact in Alzheimer's disease (AD), but there is controversy about its relevance at the early stages of the disease and in mild cognitive impairment (MCI). In vivo positron emission tomography imaging of cortical acetylcholine esterase (AChE) activity as a marker of cholinergic innervation that is expressed by cholinergic axons and cholinoceptive neurons has demonstrated a reduction of this enzyme activity in manifest AD. The technique is also useful to measure the inhibition of cerebral AChE induced by cholinesterase inhibitors for treatment of dementia symptoms. A reduction of cortical AchE activity was found consistently in all studies of AD and in few cases of MCI who later concerted to AD. The in vivo findings in MCI and very mild AD are still preliminary, and studies seem to suggest that cholinergic innervation and AChE as the main degrading enzyme are both reduced, which might result in partial compensation of their effect. (orig.)

  19. Covalent immobilization of cholesterol esterase and cholesterol oxidase on polyaniline films for application to cholesterol biosensor

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Suman [Biomolecular Electronics and Conducting Polymer Research Group, National Physical Laboratory, Dr. K. S. Krishnan Marg, New Delhi-110012 (India); Solanki, Pratima R. [Biomolecular Electronics and Conducting Polymer Research Group, National Physical Laboratory, Dr. K. S. Krishnan Marg, New Delhi-110012 (India); Pandey, M.K. [Biomolecular Electronics and Conducting Polymer Research Group, National Physical Laboratory, Dr. K. S. Krishnan Marg, New Delhi-110012 (India); Malhotra, B.D. [Biomolecular Electronics and Conducting Polymer Research Group, National Physical Laboratory, Dr. K. S. Krishnan Marg, New Delhi-110012 (India)]. E-mail: bansi@mail.nplindia.ernet.in

    2006-05-24

    Cholesterol esterase (ChEt) and cholesterol oxidase (ChOx) have been covalently immobilized on electrochemically prepared polyaniline (PANI) films. These PANI/ChEt/ChOx enzyme films have been characterized using UV-visible, Fourier transform infrared (FTIR) spectroscopy and scanning electron microscopy (SEM). Electrochemical behavior of these films has been studied using cyclic voltammetry (CV) and amperometric techniques, respectively. The PANI/ChEt/ChOx enzyme films show broad oxidation peak from 0.2 to 0.5 V. These PANI/ChEt/ChOx biosensing electrodes have a response time of about 40 s, linearity from 50 to 500 mg/dl of cholesterol oleate concentration. These PANI/ChEt/ChOx films are thermally stable up to 46 deg. C. This polyaniline based cholesterol biosensor has optimum pH in the range of 6.5-7.5, sensitivity as 7.5 x 10{sup -4} nA/mg dl and a lifetime of about 6 weeks.

  20. Pectin methyl esterases and pectins in normal and hyperhydric shoots of carnation cultured in vitro.

    Science.gov (United States)

    Saher, Shady; Piqueras, Abel; Hellin, Eladio; Olmos, Enrique

    2005-02-01

    Control and hyperhydric micropropagated plantlets from three carnation cultivars have been used to study their pectin composition and the activity of pectin methyl esterases (PMEs; EC 3.1.1.11). Pectins are a highly heterogeneous group of polymers that contribute to cell adhesion, cell wall architecture, and cell wall mechanical strength. Pectins control cell wall porosity and cell wall ionic status and are implicated in intercellular space development. The degree of esterification of pectins is controlled by the activity of cell wall PMEs; their different actions can affect the properties of the cell wall, which have been considered important with respect to controlling the development of hyperhydricity. The total pectins of hyperhydric leaves of the three varieties were significantly reduced in comparison with controls. The pectate fraction was significantly increased in hyperhydric leaves of all varieties while soluble pectins and protopectins were significantly lower. The PME activity of hyperhydric leaves was higher (4-10 times) compared to controls of the three varieties. Isoelectric focusing of PME isozymes revealed the presence of three isoforms; neutral PME activity was the major isozyme in control and hyperhydric leaves of the three varieties, whilst a decrease in the activity of the acidic isoforms was observed in hyperhydric leaves. The different PME activities could regulate some of the structural changes related to hyperhydricity in micropropagated carnation plants.

  1. [Anaesthesic management of vaginal delivery in a parturient with C1 esterase deficiency].

    Science.gov (United States)

    Libert, N; Schérier, S; Dubost, C; Franck, L; Rouquette, I; Tortosa, J-C; Rousseau, J-M

    2009-04-01

    Hereditary and acquired angioedema (HAE/AAE) are the clinical translation of a qualitative or a quantitative deficit of C1 esterase inhibitor (C1 INH). The frequency and severity of clinical manifestations vary greatly, ranging from a moderate swelling of the extremities to obstruction of upper airway. Anaesthesiologists and intensivists must be prepared to manage acute manifestations of this disease in case of life-threatening laryngeal edema. Surgery, physical trauma and labour are classical triggers of the disease. The anaesthesiologists should be aware of the drugs used as prophylaxis and treatment of acute attacks when considering labour and caesarean section. Androgens are contraindicated during pregnancy. If prophylaxis is required, tranexamic acid may be used with caution. The safest obstetric approach appears to be to administer a predelivery infusion of C1 INH concentrate. It is important to avoid manipulation of the airway as much as possible by relying on regional techniques. We report the case of a patient suffering from an HAE discovered during pregnancy. The management included administration of C1 INH during labor and early epidural analgesia for pain relief. A short review of the pathophysiology and therapeutic options follows.

  2. Characterization and purification of a bacterial chlorogenic acid esterase detected during the extraction of chlorogenic acid from arbuscular mycorrhizal tomato roots.

    Science.gov (United States)

    Negrel, Jonathan; Javelle, Francine; Morandi, Dominique; Lucchi, Géraldine

    2016-12-01

    A Gram-negative bacterium able to grow using chlorogenic acid (5-caffeoylquinic acid) as sole carbon source has been isolated from the roots of tomato plants inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis. An intracellular esterase exhibiting very high affinity (K m  = 2 μM) for chlorogenic acid has been extracted and purified by FPLC from the chlorogenate-grown cultures of this bacterium. The molecular mass of the purified esterase determined by SDS-PAGE was 61 kDa and its isoelectric point determined by chromatofocusing was 7.75. The esterase hydrolysed chlorogenic acid analogues (caffeoylshikimate, and the 4- and 3-caffeoylquinic acid isomers), feruloyl esterases substrates (methyl caffeate and methyl ferulate), and even caffeoyl-CoA in vitro but all of them were less active than chlorogenic acid, demonstrating that the esterase is a genuine chlorogenic acid esterase. It was also induced when the bacterial strain was cultured in the presence of hydroxycinnamic acids (caffeic, p-coumaric or ferulic acid) as sole carbon source, but not in the presence of simple phenolics such as catechol or protocatechuic acid, nor in the presence of organic acids such as succinic or quinic acids. The purified esterase was remarkably stable in the presence of methanol, rapid formation of methyl caffeate occurring when its activity was measured in aqueous solutions containing 10-60% methanol. Our results therefore show that this bacterial chlorogenase can catalyse the transesterification reaction previously detected during the methanolic extraction of chlorogenic acid from arbuscular mycorrhizal tomato roots. Data are presented suggesting that colonisation by Rhizophagus irregularis could increase chlorogenic acid exudation from tomato roots, especially in nutrient-deprived plants, and thus favour the growth of chlorogenate-metabolizing bacteria on the root surface or in the mycorhizosphere. Copyright © 2016 Elsevier Masson SAS. All rights

  3. PRODUCTION AND CHARACTERIZATION OF AN ALKALOTHERMOSTABLE, ORGANIC SOLVENT TOLERANT AND SURFACTANT TOLERANT ESTERASE PRODUCED BY A THERMOPHILIC BACTERIUM GEOBACILLUS SP. AGP-04, ISOLATED FROM BAKRESHWAR HOT SPRING, INDIA

    Directory of Open Access Journals (Sweden)

    Amit Ghati

    2013-10-01

    Full Text Available A thermophilic bacteria, Geobacillus sp. AGP-04, isolated from Surya Kund hot spring, Bakreshwar, West Bengal, India was studied in terms of capability of tributyrin hydrolysis and characterization of its thermostable esterase activity using p-nitrophenyl butyrate (PNPB as substrate. The extracellular crude preparation was characterized in terms of pH and temperature optima and stability, organic solvent tolerance capacity and stability, substrate specificity, surfactant tolerance capacity, kinetic parameters and activation/inhibition behavior towards some metal ions and chemicals. Tributyrin agar assay exhibited that Geobacillus sp. AGP-04 secretes an extracellular esterase. The Vmax and Km values of the esterase were found to be 5099 U/Land 103.5µM, respectively in the presence of PNPB as substrate. The optimum temperature and pH, for Geobacillus sp. AGP-04 esterase was 60oC and 8.0, respectively. Although the enzyme activity was not significantly altered by incubating crude extract solution at 20-70oC for 1 hour, the enzyme activity was fully lost at 90oC for same incubation period. The pH stability profile showed that original crude esterase activity is stable at a broad range (pH 5.0-10.0. Moreover, the enzyme was highly organic solvent and surfactant tolerant. The effect of some chemical on crude esterase activity indicated that Geobacillus sp. AGP-04 produce an esterase which contains a serine residue in active site and for its activity -SH groups are essential. Besides, enzyme production was highly induced if fermentation medium contain polysaccharides and oil as carbon source.

  4. (e,3e) collisions on He in the quasi-dipolar regime

    International Nuclear Information System (INIS)

    Taouil, I.; Lahmam-Bennani, A.; Duguet, A.

    1999-01-01

    We report the first measurement of the angular distributions of the two ejected electrons following electron impact double ionization of helium, using a multi-parameter (e,3e) spectrometer. The high incident energy of these experiments allows comparison with photo-double ionization results. Comparison is also made with existing (e,3e) theoretical calculations. (orig.)

  5. PXR agonism decreases plasma HDL levels in ApoE*3-Leiden.CETP mice

    NARCIS (Netherlands)

    Haan, W. de; Vries-van der Weij, J. de; Mol, I.M.; Hoekstra, M.; Romijn, J.A.; Jukema, J.W.; Havekes, L.M.; Princen, H.M.G.; Rensen, P.C.N.

    2009-01-01

    Pregnane X receptor (PXR) agonism has been shown to affect multiple steps in both the synthesis and catabolism of HDL, but its integrated effect on HDL metabolism in vivo remains unclear. The aim of this study was to evaluate the net effect of PXR agonism on HDL metabolism in ApoE*3-Leiden (E3L) and

  6. Procedure Redesign Methods : E3-Control: a redesign methodology for control procedures

    NARCIS (Netherlands)

    Liu, J.; Hofman, W.J.; Tan, Y.H.

    2011-01-01

    This chapter highlights the core research methodology, e3-control, that is applied throughout the ITAIDE project for the purpose of control procedure redesign. We present the key concept of the e3-control methodology and its technical guidelines. Based on the output of this chapter, domain experts

  7. 17 CFR 270.22e-3 - Exemption for liquidation of money market funds.

    Science.gov (United States)

    2010-04-01

    ... money market funds. 270.22e-3 Section 270.22e-3 Commodity and Securities Exchanges SECURITIES AND... for liquidation of money market funds. (a) Exemption. A registered open-end management investment company or series thereof (“fund”) that is regulated as a money market fund under § 270.2a-7 is exempt...

  8. E3 ubiquitin ligases as drug targets and prognostic biomarkers in melanoma

    Directory of Open Access Journals (Sweden)

    Kristina Bielskienė

    2015-01-01

    E3 ligases are of interest as drug targets for their ability to regulate proteins stability and functions. Compared to the general proteasome inhibitor bortezomib, which blocks the entire protein degradation, drugs that target a particular E3 ligase are expected to have better selectivity with less associated toxicity. Components of different E3 ligases complexes (FBW7, MDM2, RBX1/ROC1, RBX2/ROC2, cullins and many others are known as oncogenes or tumor suppressors in melanomagenesis. These proteins participate in regulation of different cellular pathways and such important proteins in cancer development as p53 and Notch. In this review we summarized published data on the role of known E3 ligases in the development of melanoma and discuss the inhibitors of E3 ligases as a novel approach for the treatment of malignant melanomas.

  9. Avoidance behaviour response and esterase inhibition in the earthworm, Lumbricus terrestris, after exposure to chlorpyrifos.

    Science.gov (United States)

    Martínez Morcillo, S; Yela, J L; Capowiez, Y; Mazzia, C; Rault, M; Sanchez-Hernandez, Juan C

    2013-05-01

    The avoidance response of earthworms to polluted soils has been standardised using a simple and low-cost test, which facilitates soil toxicity screening. In this study, the avoidance response of Lumbricus terrestris was quantified in chlorpyrifos-spiked soils, depending on the pesticide concentration and exposure duration. The inhibition of acetylcholinesterase (AChE) and carboxylesterase (CbE) activities was also determined as indirect measures of pesticide bioavailability. The effects of different chlorpyrifos concentrations were examined in a standardised test (two-chamber system) with 0.6, 3 and 15 mg/kg chlorpyrifos. A modification of the test involved a pre-exposure step (24, 48 or 72 h) in soils spiked with 15 mg/kg. In both protocols, earthworms were unable to avoid the contaminated soils. However, the esterase activities showed that all earthworms were exposed to chlorpyrifos. Acetylcholinesterase activity did not change in earthworms in the standardised behavioural test (0.58 ± 0.20 U/mg protein, mean ± SD; n = 72), whereas the CbE activity was significantly inhibited (62-87 % inhibition) in earthworms exposed to 3 and 15 mg/kg. In the modified test, earthworms had greatly inhibited AChE activity (0.088 ± 0.034 U/mg protein, n = 72), which was supported by reactivation of the inhibited enzyme activity in the presence of pralidoxime (2-PAM). Similarly, the CbE activity was significantly inhibited in earthworms with all treatments. This study suggests that the avoidance behaviour test for organophosphorus-contaminated soils could be supported by specific biomarkers to facilitate a better understanding of pesticide exposure and toxicity during this test.

  10. Functional tuning of the catalytic residue pKa in a de novo designed esterase.

    Science.gov (United States)

    Hiebler, Katharina; Lengyel, Zsófia; Castañeda, Carlos A; Makhlynets, Olga V

    2017-09-01

    AlleyCatE is a de novo designed esterase that can be allosterically regulated by calcium ions. This artificial enzyme has been shown to hydrolyze p-nitrophenyl acetate (pNPA) and 4-nitrophenyl-(2-phenyl)-propanoate (pNPP) with high catalytic efficiency. AlleyCatE was created by introducing a single-histidine residue (His 144 ) into a hydrophobic pocket of calmodulin. In this work, we explore the determinants of catalytic properties of AlleyCatE. We obtained the pK a value of the catalytic histidine using experimental measurements by NMR and pH rate profile and compared these values to those predicted from electrostatics pK a calculations (from both empirical and continuum electrostatics calculations). Surprisingly, the pK a value of the catalytic histidine inside the hydrophobic pocket of calmodulin is elevated as compared to the model compound pK a value of this residue in water. We determined that a short-range favorable interaction with Glu 127 contributes to the elevated pK a of His 144 . We have rationally modulated local electrostatic potential in AlleyCatE to decrease the pK a of its active nucleophile, His 144 , by 0.7 units. As a direct result of the decrease in the His 144 pK a value, catalytic efficiency of the enzyme increased by 45% at pH 6. This work shows that a series of simple NMR experiments that can be performed using low field spectrometers, combined with straightforward computational analysis, provide rapid and accurate guidance to rationally improve catalytic efficiency of histidine-promoted catalysis. Proteins 2017; 85:1656-1665. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  11. A stereospecific carboxyl esterase from Bacillus coagulans hosting nonlipase activity within a lipase-like fold.

    Science.gov (United States)

    De Vitis, Valerio; Nakhnoukh, Cristina; Pinto, Andrea; Contente, Martina L; Barbiroli, Alberto; Milani, Mario; Bolognesi, Martino; Molinari, Francesco; Gourlay, Louise J; Romano, Diego

    2018-03-01

    Microbial carboxylesterases are important biocatalysts that selectively hydrolyze an extensive range of esters. Here, we report the biochemical and structural characterization of an atypical carboxylesterase from Bacillus coagulans (BCE), endowed with high enantioselectivity toward different 1,2-O-isopropylideneglycerol (IPG or solketal) esters. BCE efficiently catalyzes the production of enantiopure (S)-IPG, a chiral building block for the synthesis of β-blockers, glycerophospholipids, and prostaglandins; efficient hydrolysis was observed up to 65 °C. To gain insight into the mechanistic bases of such enantioselectivity, we solved the crystal structures of BCE in apo- and glycerol-bound forms at resolutions of 1.9 and 1.8 Å, respectively. In silico docking studies on the BCE structure confirmed that IPG esters with small acyl chains (≤ C6) were easily accommodated in the active site pocket, indicating that small conformational changes are necessary to accept longer substrates. Furthermore, docking studies suggested that enantioselectivity may be due to an improved stabilization of the tetrahedral reaction intermediate for the S-enantiomer. Contrary to the above functional data implying nonlipolytic functions, BCE displays a lipase-like 3D structure that hosts a "lid" domain capping the main entrance to the active site. In lipases the lid mediates catalysis through interfacial activation, a process that we did not observe for BCE. Overall, we present the functional-structural properties of an atypical carboxyl esterase that has nonlipase-like functions, yet possesses a lipase-like 3D fold. Our data provide original enzymatic information in view of BCE applications as an inexpensive, efficient biocatalyst for the production of enantiopure (S)-IPG. Coordinates and structure factors have been deposited in the Protein Data Bank (www.rcsb.org) under accession numbers 5O7G (apo-BCE) and 5OLU (glycerol-bound BCE). © 2017 Federation of European Biochemical

  12. Vaginal Fornix Discharge Cellularity and Its Leukocyte Esterase Activity for Diagnosis of Endometritis in Dairy Cows

    Directory of Open Access Journals (Sweden)

    Abolfazl HAJIBEMANI

    2016-01-01

    Full Text Available The objective of the present study was to evaluate the application of some strip test markers (i.e., leukocyte esterase (LE activity, protein, nitrate and pH for diagnosis of endometritis in dairy cows using vaginal fornix discharge. Also, the total white blood cell count (t-WBC/l of this secretion and degenerative changes of neutrophils in cervical cytology were used as alternative methods to predict progression of the endometritis severity. Holstein cows (n=215 between 30-40 days in milk (DIM were included and examined. Giemsa-stained smear was prepared from cervical mucus. Cervical cytology test was considered as reference screening method for the detection of subclinical endometritis. The LE activity and t-WBC in the vaginal fornix discharge of subclinical endometritis cows were significantly higher than those from healthy cows. Sensitivity and specificity were 78% and 73% for LE10 activity (10 minutes after contacting with discharges and 60% and 69% for t-WBC (cut off point=210 cells/l for diagnosis of subclinical endometritis, respectively. There was a good agreement between LE10 activity, t-WBC and cervical cytology test with a Kappa coefficient of 0.4 and 0.42, respectively (P<0.0001. Total WBC count in discharge and degenerative neutrophils (DN percentages increase simultaneously with the degree and severity of endometritis. There was a highly significant (P<0.01 correlation between t-WBC and some reagent strip test markers (LE activity, protein and nitrate in clear discharge of studied cows. In conclusion, the present results suggest the LE activity and t-WBC in vaginal fornix discharge could be used as non-invasive reliable and valid methods for screening of subclinical endometritis in postpartum dairy herds.

  13. Immobilization of cholesterol esterase in mesoporous silica materials and its hydrolytic activity toward diethyl phthalate

    Energy Technology Data Exchange (ETDEWEB)

    Orita, Toru, E-mail: nqj45366@nifty.com [Division of Chemistry for Materials, Graduate School of Engineering, Mie University, 1577 Kurimamachiya-cho, Tsu, Mie 514-8570 (Japan); Taiyo Kagaku Co. Ltd., 800 Yamada-cho, Yokkaichi, Mie 512-1111 (Japan); Tomita, Masahiro [Division of Chemistry for Materials, Graduate School of Engineering, Mie University, 1577 Kurimamachiya-cho, Tsu, Mie 514-8570 (Japan); Saito, Takao; Nishida, Nasakazu; Kato, Katsuya [National Institute of Advanced Industrial Science and Technology, 2266-78 Anagahora, Moriyamaku, Nagoya, Aichi 463-8560 (Japan)

    2012-05-01

    Cholesterol esterase (CE, cholesteryl ester hydrolase, EC 3.1.1.13) from porcine pancreas (molecular weight 400-500 kDa) exhibits hydrolytic activity toward various toxic organic phthalate esters. CE was confined in the nanospace (diameter 3-30 nm) of five types of mesoporous silica (MPS) that differ in structural properties such as pore diameter, pore volume, and particle morphology. These structural properties were characterized by transmission electron microscopy, small-angle X-ray diffraction, N{sub 2} adsorption-desorption experiments, solid-state {sup 13}C nuclear magnetic resonance (NMR), and solid-state {sup 29}Si NMR. Catalytic activities of immobilized and free CE were evaluated by the hydrolysis of diethyl phthalate in phosphate buffer solutions containing an organic cosolvent. Optimal activity recovery was achieved when CE was immobilized in n-decane-functionalized MPS, which had a large pore size (22.5 nm). The immobilization also protected against effects of temperature within the range 30 Degree-Sign C-60 Degree-Sign C; CE immobilized in n-decyl-functionalized MPS exhibited better thermal stability than in non-functionalized MPS or free CE. Moreover, it retained approximately 60% of its catalytic activity even after six catalytic cycles. - Highlights: Black-Right-Pointing-Pointer The highest activity of immobilized CE was shown in MPS with a pore size of 22.5 nm. Black-Right-Pointing-Pointer Catalytic efficiency improved when MPS was functionalized by n-decyl substitution. Black-Right-Pointing-Pointer Immobilized CE exhibited good thermal stability and reusability. Black-Right-Pointing-Pointer Organic co-solvent and the substrate structures affected enzyme activities.

  14. Crystal structure of hyperthermophilic esterase EstE1 and the relationship between its dimerization and thermostability properties

    Directory of Open Access Journals (Sweden)

    Koh Eunhee

    2007-07-01

    Full Text Available Abstract Background EstE1 is a hyperthermophilic esterase belonging to the hormone-sensitive lipase family and was originally isolated by functional screening of a metagenomic library constructed from a thermal environmental sample. Dimers and oligomers may have been evolutionally selected in thermophiles because intersubunit interactions can confer thermostability on the proteins. The molecular mechanisms of thermostabilization of this extremely thermostable esterase are not well understood due to the lack of structural information. Results Here we report for the first time the 2.1-Å resolution crystal structure of EstE1. The three-dimensional structure of EstE1 exhibits a classic α/β hydrolase fold with a central parallel-stranded beta sheet surrounded by alpha helices on both sides. The residues Ser154, Asp251, and His281 form the catalytic triad motif commonly found in other α/β hydrolases. EstE1 exists as a dimer that is formed by hydrophobic interactions and salt bridges. Circular dichroism spectroscopy and heat inactivation kinetic analysis of EstE1 mutants, which were generated by structure-based site-directed mutagenesis of amino acid residues participating in EstE1 dimerization, revealed that hydrophobic interactions through Val274 and Phe276 on the β8 strand of each monomer play a major role in the dimerization of EstE1. In contrast, the intermolecular salt bridges contribute less significantly to the dimerization and thermostability of EstE1. Conclusion Our results suggest that intermolecular hydrophobic interactions are essential for the hyperthermostability of EstE1. The molecular mechanism that allows EstE1 to endure high temperature will provide guideline for rational design of a thermostable esterase/lipase using the lipolytic enzymes showing structural similarity to EstE1.

  15. The Alpha-Defensin Immunoassay and Leukocyte Esterase Colorimetric Strip Test for the Diagnosis of Periprosthetic Infection

    Science.gov (United States)

    Wyatt, M.C.; Beswick, A.D.; Kunutsor, S.K.; Wilson, M.J.; Whitehouse, M.R.; Blom, A.W.

    2016-01-01

    Background: Synovial biomarkers have recently been adopted as diagnostic tools for periprosthetic joint infection (PJI), but their utility is uncertain. The purpose of this systematic review and meta-analysis was to synthesize the evidence on the accuracy of the alpha-defensin immunoassay and leukocyte esterase colorimetric strip test for the diagnosis of PJI compared with the Musculoskeletal Infection Society diagnostic criteria. Methods: We performed a systematic review to identify diagnostic technique studies evaluating the accuracy of alpha-defensin or leukocyte esterase in the diagnosis of PJI. MEDLINE and Embase on Ovid, ACM, ADS, arXiv, CERN DS (Conseil Européen pour la Recherche Nucléaire Document Server), CrossRef DOI (Digital Object Identifier), DBLP (Digital Bibliography & Library Project), Espacenet, Google Scholar, Gutenberg, HighWire, IEEE Xplore (Institute of Electrical and Electronics Engineers digital library), INSPIRE, JSTOR (Journal Storage), OAlster (Open Archives Initiative Protocol for Metadata Harvesting), Open Content, Pubget, PubMed, and Web of Science were searched for appropriate studies indexed from inception until May 30, 2015, along with unpublished or gray literature. The classification of studies and data extraction were performed independently by 2 reviewers. Data extraction permitted meta-analysis of sensitivity and specificity with construction of receiver operating characteristic curves for each test. Results: We included 11 eligible studies. The pooled diagnostic sensitivity and specificity of alpha-defensin (6 studies) for PJI were 1.00 (95% confidence interval [CI], 0.82 to 1.00) and 0.96 (95% CI, 0.89 to 0.99), respectively. The area under the curve (AUC) for alpha-defensin and PJI was 0.99 (95% CI, 0.98 to 1.00). The pooled diagnostic sensitivity and specificity of leukocyte esterase (5 studies) for PJI were 0.81 (95% CI, 0.49 to 0.95) and 0.97 (95% CI, 0.82 to 0.99), respectively. The AUC for leukocyte esterase and PJI

  16. Comparative Study of Esterase and Hemolytic Activities in Clinically Important Candida Species, Isolated From Oral Cavity of Diabetic and Non-diabetic Individuals.

    Science.gov (United States)

    Fatahinia, Mahnaz; Poormohamadi, Farzad; Zarei Mahmoudabadi, Ali

    2015-03-01

    Diabetes mellitus as a chronic metabolic disease occurs in patients with partial or complete deficiency of insulin secretion or disorder in action of insulin on tissue. The disease is known to provide conditions for overgrowth of Candida species. Candida spp. cause candidiasis by many virulence factors such as esterase, hemolysin and phospholipase. This study aimed to compare esterase and hemolytic activity in various Candida species isolated from oral cavity of diabetic and non-diabetic individuals. Swab samples were taken from 95 patients with diabetes (35 men and 60 women) and 95 normal persons (42 men and 53 women) and cultured on Sabouraud dextrose agar. Identification of isolated yeasts was performed by germ tube test, morphology on CHROMagar Candida medium, corn meal agar and ability to grow at 45°C. Hemolysin activity was evaluated using blood plate assay and esterase activity was determined using the Tween 80 opacity test. Different Candida species were isolated from 57 (60%) diabetic and 24 (25%) non-diabetic individuals. Esterase activity was detected in all Candida isolates. Only 21.6% of C. albicans from patients with diabetes had esterase activity as + 3, while it ranged from + 1 to + 2 in others. Hemolytic activity was determined in C. albicans, C. dubliniensis, C. glabrata and C. krusei as 0.79, 0.58, 0.66 and 0.74, respectively. Hemolytic activity was significantly different in the two groups of diabetics and non-diabetics. Oral carriage of C. albicans in the diabetic group (n = 42; 66.7%) was significantly greater than the control group (n = 16; 57.1%). Esterase activity of C. albicans in diabetic group was higher than non-diabetic group. Although C. albicans remains the most frequently pathogenic yeast for human, but other species are increasing.

  17. First results with the general equilibrium model GEM-E3 Switzerland

    International Nuclear Information System (INIS)

    Bahn, O.; Frei, C.

    2000-01-01

    The GEM-E3 model has been implemented and applied for Switzerland. It has been in particular used to assess an ecological tax reform in Switzerland. Results of this analysis are presented here. (author)

  18. Scaling of cross-sections for asymmetric (e,3e) process on helium ...

    Indian Academy of Sciences (India)

    , India ... (e, 3e) process; five-fold differential cross-section; scaling; helium isoelec- tronic ions. ... ration and experimental control of the target and the intensity related problems make the measurements extremely difficult. The scaling laws of ...

  19. Redefining E-3 Core Competencies for Dominant Battlespace Knowledge in Future Combat Operations

    National Research Council Canada - National Science Library

    Kirkendall, David A

    2005-01-01

    .... The study focuses on how E-3 training is driven by the maintenance of a set of battle management core competencies rooted in the basics of aircraft tactical fluid control force accountability and aerial refueling...

  20. E3 Success Story - Whirlpool Trains Staff on Lean and Green Advantage

    Science.gov (United States)

    Whirlpool Corporation invited Green Suppliers Network representatives to its Monterrey facility to provide training on the Lean and Green Advantage. The project sought to expand E3 initiatives to every part of the company's operations.

  1. E3 Success Story - Advancing Performance in Sustainability and Workforce Development

    Science.gov (United States)

    E3: North Carolina advances performance in sustainability and workforce development strategies for the state's manufacturers. The initiative helps communities and manufacturers address energy and sustainability challenges by leveraging expertise.

  2. A study of the profile of the E3 electron trap in GaAs

    Energy Technology Data Exchange (ETDEWEB)

    Kourkoutas, C.D. (TEI Athens (Greece). Dept. of Physics Chemistry and Material Technology); Kovacs, B.; Szentpali, B.; Somogyi, K. (Research Inst. for Technical Physics, Budapest (Hungary)); Euthymiou, P.C. (Athens Univ. (Greece)); Giakoumakis, G.E. (Ioannina Univ. (Greece). Dept. of Physics)

    1994-01-01

    Electron irradiation at room temperature introduces in GaAs a donor type electronic state Tx at 0.18 eV, which is associated with the E3 electron trap. The presence of Tx is observed at depths d > 1.5 [mu]m, which correspond to the limits of the depletion region under the highest applied reverse bias voltage, while the E3 trap concentration drops off into the same region. (author).

  3. A study of the profile of the E3 electron trap in GaAs

    International Nuclear Information System (INIS)

    Kourkoutas, C.D.; Euthymiou, P.C.; Giakoumakis, G.E.

    1994-01-01

    Electron irradiation at room temperature introduces in GaAs a donor type electronic state Tx at 0.18 eV, which is associated with the E3 electron trap. The presence of Tx is observed at depths d > 1.5 μm, which correspond to the limits of the depletion region under the highest applied reverse bias voltage, while the E3 trap concentration drops off into the same region. (author)

  4. Isolation and Expression analysis of OsPME1, encoding for a putative Pectin Methyl Esterase from Oryza sativa (subsp. indica)

    OpenAIRE

    Kanneganti, Vydehi; Gupta, Aditya Kumar

    2009-01-01

    Pectin Methyl Esterases (PMEs) play an essential role during plant development by affecting the mechanical properties of the plant cell walls. Recent studies indicated that PMEs play important role in pollen tube development. In this study, we isolated a 1.3 kb cDNA clone from rice panicle cDNA library. It contained a 1038 bp of open reading frame (ORF) encoding for a putative pectin methyl esterase of 345 aminoacids with a 20 aminoacid signal peptide and was hence designated as OsPME1 (Oryza...

  5. The Use of Plasma-Derived Complement C1-Esterase Inhibitor Concentrate (Berinert®) in the Treatment of Angiotensin Converting Enzyme-Inhibitor Related Angioedema

    DEFF Research Database (Denmark)

    Hermanrud, Thorbjørn; Duus, Nicolaj; Bygum, Anette

    2016-01-01

    Angioedema of the upper airways is a severe and potentially life-threatening condition. The incidence has been increasing in the past two decades, primarily due to pharmaceuticals influencing the generation or degradation of the vasoactive molecule bradykinin. Plasma-derived C1-esterase inhibitor...... concentrate is a well-established treatment option of hereditary and acquired complement C1-esterase inhibitor deficiency, which are also mediated by an increased level of bradykinin resulting in recurrent angioedema. We here present a case of severe angiotensin converting enzyme-inhibitor related angioedema...

  6. Prevalence of positive urinary dipstick analysis (leucocyte esterase, nitrite, haemoglobin, or glucose) in a population of 3645 adult subjects--consequence for measurement of urinary albumin excretion rate

    DEFF Research Database (Denmark)

    Clausen, P; Jensen, J S; Borch-Johnsen, K

    1998-01-01

    OBJECTIVES: To assess prevalence of positive urinary dipstick analysis for leucocyte esterase, nitrite, haemoglobin, or glucose in the general population and measure the urinary albumin excretion rate (UAER) in subjects with or without a positive dipstick analysis. DESIGN: A cross-sectional study...... of 3645 subjects. SETTING: An unselected urban population study. MAIN OUTCOME MEASURES: Prevalence data of positive dipstick analyses and UAER values. RESULTS: Prevalence data of a positive dipstick analysis were 12%, 4%, 3% and 6%, respectively, for leucocyte esterase, nitrite, haemoglobin, and glucose...

  7. Molecular structure and spectral properties of ethyl 3-quinolinecarboxylate (E3Q) and [Ag(E3Q)2(TCA)] complex (TCA = Trichloroacetate)

    Science.gov (United States)

    Soliman, Saied M.; Kassem, Taher S.; Badr, Ahmed M. A.; Abou Youssef, Morsy A.; Assem, Rania

    2014-09-01

    A new [Ag(E3Q)2(TCA)] complex; (E3Q = Ethyl 3-quinolinecarboxylate and TCA = Trichloroacetate) has been synthesized and characterized using elemental analysis, FTIR, NMR and mass spectroscopy. The molecular geometry and spectroscopic properties of the complex as well as the free ligand have been calculated using the hybrid B3LYP method. The calculations predicted a distorted tetrahedral arrangement around Ag(I) ion. The vibrational spectra of the studied compounds have been assigned using potential energy distribution (PED). TD-DFT method was used to predict the electronic absorption spectra. The most intense absorption band showed a bathochromic shift and lowering of intensity in case of the complex (233.7 nm, f = 0.5604) compared to E3Q (λmax = 228.0 nm, f = 0.9072). The calculated 1H NMR chemical shifts using GIAO method showed good correlations with the experimental data. The computed dipole moment, polarizability and HOMO-LUMO energy gap were used to predict the nonlinear optical (NLO) properties. It is found that Ag(I) enhances the NLO activity. The natural bond orbital (NBO) analyses were used to elucidate the intramolecular charge transfer interactions causing stabilization for the investigated systems.

  8. Loss of the E3 ubiquitin ligase LRSAM1 sensitizes peripheral axons to degeneration in a mouse model of Charcot-Marie-Tooth disease

    OpenAIRE

    Bogdanik, Laurent P.; Sleigh, James N.; Tian, Cong; Samuels, Mark E.; Bedard, Karen; Seburn, Kevin L.; Burgess, Robert W.

    2013-01-01

    SUMMARY Charcot-Marie-Tooth disease (CMT) is a clinically and genetically heterogeneous condition characterized by peripheral axon degeneration with subsequent motor and sensory deficits. Several CMT gene products function in endosomal sorting and trafficking to the lysosome, suggesting that defects in this cellular pathway might present a common pathogenic mechanism for these conditions. LRSAM1 is an E3 ubiquitin ligase that is implicated in this process, and mutations in LRSAM1 have rece...

  9. Crystallization and preliminary X-ray analysis of a novel esterase Rv0045c from Mycobacterium tuberculosis

    International Nuclear Information System (INIS)

    Xu, Lipeng; Guo, Jiubiao; Zheng, Xiangdong; Wen, Tingyi; Sun, Fei; Liu, Siguo; Pang, Hai

    2010-01-01

    The novel esterase Rv0045c from M. tuberculosis was expressed and purified to homogeneity. The crystals of native and SeMet-labelled Rv0045c protein that were obtained diffracted to resolutions of 2.7 and 3.0 Å, respectively. The Rv0045c protein is predicted to be an esterase that is involved in lipid metabolism in Mycobacterium tuberculosis. The protein was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. The Rv0045c protein crystals diffracted to a resolution of 2.7 Å using a synchrotron-radiation source and belonged to space group P3 1 or P3 2 , with unit-cell parameters a = b = 73.465, c = 48.064 Å, α = β = 90, γ = 120°. Purified SeMet-labelled Rv0045c protein was also crystallized and formed crystals that diffracted to a resolution of 3.0 Å using an in-house X-ray radiation source

  10. Inhibition, recovery and oxime-induced reactivation of muscle esterases following chlorpyrifos exposure in the earthworm Lumbricus terrestris

    International Nuclear Information System (INIS)

    Collange, B.; Wheelock, C.E.; Rault, M.; Mazzia, C.; Capowiez, Y.; Sanchez-Hernandez, J.C.

    2010-01-01

    Assessment of wildlife exposure to organophosphorus (OP) pesticides generally involves the measurement of cholinesterase (ChE) inhibition, and complementary biomarkers (or related endpoints) are rarely included. Herein, we investigated the time course inhibition and recovery of ChE and carboxylesterase (CE) activities in the earthworm Lumbricus terrestris exposed to chlorpyrifos, and the ability of oximes to reactivate the phosphorylated ChE activity. Results indicated that these esterase activities are a suitable multibiomarker scheme for monitoring OP exposure due to their high sensitivity to OP inhibition and slow recovery to full activity levels following pesticide exposure. Moreover, oximes reactivated the inhibited ChE activity of the earthworms exposed to 12 and 48 mg kg -1 chlorpyrifos during the first week following pesticide exposure. This methodology is useful for providing evidence for OP-mediated ChE inhibition in individuals with a short history of OP exposure (≤1 week); resulting a valuable approach for assessing multiple OP exposure episodes in the field. - Esterase inhibition combined with oxime reactivation methods is a suitable approach for monitoring organophosphate contamination

  11. Inhibition, recovery and oxime-induced reactivation of muscle esterases following chlorpyrifos exposure in the earthworm Lumbricus terrestris

    Energy Technology Data Exchange (ETDEWEB)

    Collange, B. [Universite d' Avignon et des Pays de Vaucluse, UMR 406 Abeilles et Environnement, Site AGROPARC, F-84914, Avignon Cede 09 (France); Wheelock, C.E. [Division of Physiological Chemistry II, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE 171 77, Stockholm (Sweden); Rault, M.; Mazzia, C. [Universite d' Avignon et des Pays de Vaucluse, UMR 406 Abeilles et Environnement, Site AGROPARC, F-84914, Avignon Cede 09 (France); Capowiez, Y. [INRA, Unite PSH, Site AGROPARC, F-84914 Avignon Cedex 09 (France); Sanchez-Hernandez, J.C., E-mail: juancarlos.sanchez@uclm.e [Laboratory of Ecotoxicology, Faculty of Environmental Science, University of Castilla-La Mancha, Avda. Carlos III s/n, 45071, Toledo (Spain)

    2010-06-15

    Assessment of wildlife exposure to organophosphorus (OP) pesticides generally involves the measurement of cholinesterase (ChE) inhibition, and complementary biomarkers (or related endpoints) are rarely included. Herein, we investigated the time course inhibition and recovery of ChE and carboxylesterase (CE) activities in the earthworm Lumbricus terrestris exposed to chlorpyrifos, and the ability of oximes to reactivate the phosphorylated ChE activity. Results indicated that these esterase activities are a suitable multibiomarker scheme for monitoring OP exposure due to their high sensitivity to OP inhibition and slow recovery to full activity levels following pesticide exposure. Moreover, oximes reactivated the inhibited ChE activity of the earthworms exposed to 12 and 48 mg kg{sup -1} chlorpyrifos during the first week following pesticide exposure. This methodology is useful for providing evidence for OP-mediated ChE inhibition in individuals with a short history of OP exposure (<=1 week); resulting a valuable approach for assessing multiple OP exposure episodes in the field. - Esterase inhibition combined with oxime reactivation methods is a suitable approach for monitoring organophosphate contamination

  12. Evolution of the feruloyl esterase MtFae1a from Myceliophthora thermophila towards improved catalysts for antioxidants synthesis.

    Science.gov (United States)

    Varriale, Simona; Cerullo, Gabriella; Antonopoulou, Io; Christakopoulos, Paul; Rova, Ulrika; Tron, Thierry; Fauré, Régis; Jütten, Peter; Piechot, Alexander; Brás, Joana L A; Fontes, Carlos M G A; Faraco, Vincenza

    2018-04-23

    The chemical syntheses currently employed for industrial purposes, including in the manufacture of cosmetics, present limitations such as unwanted side reactions and the need for harsh chemical reaction conditions. In order to overcome these drawbacks, novel enzymes are developed to catalyze the targeted bioconversions. In the present study, a methodology for the construction and the automated screening of evolved variants library of a Type B feruloyl esterase from Myceliophthora thermophila (MtFae1a) was developed and applied to generation of 30,000 mutants and their screening for selecting the variants with higher activity than the wild-type enzyme. The library was generated by error-prone PCR of mtfae1a cDNA and expressed in Saccharomyces cerevisiae. Screening for extracellular enzymatic activity towards 4-nitrocatechol-1-yl ferulate, a new substrate developed ad hoc for high-throughput assays of feruloyl esterases, led to the selection of 30 improved enzyme variants. The best four variants and the wild-type MtFae1a were investigated in docking experiments with hydroxycinnamic acid esters using a model of 3D structure of MtFae1a. These variants were also used as biocatalysts in transesterification reactions leading to different target products in detergentless microemulsions and showed enhanced synthetic activities, although the screening strategy had been based on improved hydrolytic activity.

  13. Heterologous production and characterization of a chlorogenic acid esterase from Ustilago maydis with a potential use in baking.

    Science.gov (United States)

    Nieter, Annabel; Kelle, Sebastian; Takenberg, Meike; Linke, Diana; Bunzel, Mirko; Popper, Lutz; Berger, Ralf G

    2016-10-15

    Ustilago maydis, an edible mushroom growing on maize (Zea mays), is consumed as the food delicacy huitlacoche in Mexico. A chlorogenic acid esterase from this basidiomycete was expressed in good yields cultivating the heterologous host Pichia pastoris on the 5L bioreactor scale (reUmChlE; 45.9UL(-1)). In contrast to previously described chlorogenic acid esterases, the reUmChlE was also active towards feruloylated saccharides. The enzyme preferred substrates with the ferulic acid esterified to the O-5 position of arabinose residues, typical of graminaceous monocots, over the O-2 position of arabinose or the O-6 position of galactose residues. Determination of kcat/Km showed that the reUmChlE hydrolyzed chlorogenic acid 18-fold more efficiently than methyl ferulate, p-coumarate or caffeate. Phenolic acids were released by reUmChlE from natural substrates, such as destarched wheat bran, sugar beet pectin and coffee pulp. Treatment of wheat dough using reUmChlE resulted in a noticeable softening indicating a potential application of the enzyme in bakery and confectionery. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Identification of Morphological Character and Esterase Isozyme Pattern in Second-Generation Black Rice Plant Irradiated to Gamma Rays

    Science.gov (United States)

    Hartanti, R. S.; Putri, T. A. N.; Zulfa, F.; Sutarno; Suranto

    2017-04-01

    Black rice is one of the functional foods due to its high anthocyanin content. Black rice grain was irradiated by gamma rays with a dose of 200 Gy and 300 Gy. The main purpose of this irradiation is to induce mutation to the black rice plant in order to achieve the improved organism. This study was undertaken to elucidate the morphological character and esterase isozyme pattern of black rice plant after irradiated by gamma rays. There were morphological differences on leaves, stems and grains between irradiated and non irradiated black rice plant. Gamma radiation dose of 200 Gy showed the significant influence of the length of the stem, number of internodes, and length of leaves. The radiation dose of 300 Gy showed the significant influence of the decrease value of diameter of 3rd internodes, number of branches and width of leaves. Flowering time is getting faster as increasing radiation dose. At the age of 74 days after planting there are 9.15% plants of 200 Gy radiation dose that have flowered faster than normal plants. This value increased into 11.45% at the dose of radiation 300 Gy. There were differences in the esterase banding pattern between radiation dose of 200 Gy and 300 Gy than the control plants, indicated that randomly mutation has occurred.

  15. Esterase- and pH-responsive poly(β-amino ester)-capped mesoporous silica nanoparticles for drug delivery

    Science.gov (United States)

    Fernando, Isurika R.; Ferris, Daniel P.; Frasconi, Marco; Malin, Dmitry; Strekalova, Elena; Yilmaz, M. Deniz; Ambrogio, Michael W.; Algaradah, Mohammed M.; Hong, Michael P.; Chen, Xinqi; Nassar, Majed S.; Botros, Youssry Y.; Cryns, Vincent L.; Stoddart, J. Fraser

    2015-04-01

    Gating of mesoporous silica nanoparticles (MSNs) with the stimuli-responsive poly(β-amino ester) has been achieved. This hybrid nanocarrier releases doxorubicin (DOX) under acidic conditions or in the presence of porcine liver esterase. The DOX loaded poly(β-amino ester)-capped MSNs reduce cell viability when tested on MDA-MB-231 human breast cancer cells.Gating of mesoporous silica nanoparticles (MSNs) with the stimuli-responsive poly(β-amino ester) has been achieved. This hybrid nanocarrier releases doxorubicin (DOX) under acidic conditions or in the presence of porcine liver esterase. The DOX loaded poly(β-amino ester)-capped MSNs reduce cell viability when tested on MDA-MB-231 human breast cancer cells. Electronic supplementary information (ESI) available: Experimental details relating to (i) the synthesis and characterisation of the surface-functionalised MSN and POL (ii) cargo-loading and release studies in solution, (iii) cellular internalisation of nanomaterials, and (iv) cell viability tests. See DOI: 10.1039/c4nr07443b

  16. In-vivo measurements of regional acetylcholine esterase activity in degenerative dementia: comparison with blood flow and glucose metabolism.

    Science.gov (United States)

    Herholz, K; Bauer, B; Wienhard, K; Kracht, L; Mielke, R; Lenz, M O; Strotmann, T; Heiss, W D

    2000-01-01

    Memory and attention are cognitive functions that depend heavily on the cholinergic system. Local activity of acetylcholine esterase (AChE) is an indicator of its integrity. Using a recently developed tracer for positron emission tomography (PET), C-11-labeled N-methyl-4-piperidyl-acetate (C11-MP4A), we measured regional AChE activity in 4 non-demented subjects, 4 patients with dementia of Alzheimer type (DAT) and 1 patient with senile dementia of Lewy body type (SDLT), and compared the findings with measurements of blood flow (CBF) and glucose metabolism (CMRGlc). Initial tracer extraction was closely related to CBF. AChE activity was reduced significantly in all brain regions in demented subjects, whereas reduction of CMRGlc and CBF was more limited to temporo-parietal association areas. AChE activity in SDLT was in the lower range of values in DAT. Our results indicate that, compared to non-demented controls, there is a global reduction of cortical AChE activity in dementia. Dementia, cholinergic system, acetylcholine esterase, positron emission tomography, cerebral blood flow, cerebral glucose metabolism.

  17. Purification and characterization of a type B feruloyl esterase (StFAE-A) from the thermophilic fungus Sporotrichum thermophile

    DEFF Research Database (Denmark)

    Topakas, E.; Stamatis, H.; Biely, P.

    2004-01-01

    A feruloyl esterase (StFAE-A) produced by Sporotrichum thermophile was purified to homogeneity. The purified homogeneous preparation of native StFAE-A exhibited a molecular mass of 57.0+/-1.5 kDa, with a mass of 33+/-1 kDa on SDS-PAGE. The pI of the enzyme was estimated by cation......-exchange chromatofocusing to be at pH 3.1. The enzyme activity was optimal at pH 6.0 and 55-60 degreesC. The purified esterase was stable at the pH range 5.0-7.0. The enzyme retained 70% of activity after 7 h at 50 degreesC and lost 50% of its activity after 45 min at 55 degreesC and after 12 min at 60 degrees......C. Determination of k(cat)/K-m revealed that the enzyme hydrolyzed methyl p-coumarate 2.5- and 12-fold more efficiently than methyl caffeate and methyl ferulate, respectively. No activity on methyl sinapinate was detected. The enzyme was active on substrates containing ferulic acid ester linked to the C-5 and C-2...

  18. Fibulin-1C, C1 Esterase Inhibitor and Glucose Regulated Protein 75 Interact with the CREC Proteins, Calumenin and Reticulocalbin.

    Directory of Open Access Journals (Sweden)

    Gry Aune Westergaard Hansen

    Full Text Available Affinity purification, immunoprecipitation, gel electrophoresis and mass spectrometry were used to identify fibulin-1C, C1 esterase inhibitor and glucose regulated protein 75, grp75, as binding partners of the CREC proteins, calumenin and reticulocalbin. Surface plasmon resonance was used to verify the interaction of all three proteins with each of the CREC proteins. Fibulin-1C interacts with calumenin and reticulocalbin with an estimated dissociation constant around 50-60 nM. The interaction, at least for reticulocalbin, was not dependent upon the presence of Ca2+. C1 esterase inhibitor interacted with both proteins with an estimated dissociation constant at 1 μM for reticulocalbin and 150 nM for calumenin. The interaction, at least for calumenin, was dependent upon the presence of Ca2+ with strong interaction at 3.5 mM while no detectable interaction could be found at 0.1 mM. Grp75 binds with an affinity of approximately 3-7 nM with reticulocalbin as well as with calumenin. These interactions suggest functional participation of the CREC proteins in chaperone activity, cell proliferation and transformation, cellular aging, haemostasis and thrombosis as well as modulation of the complement system in fighting bacterial infection.

  19. Crystallization and preliminary crystallographic analysis of an esterase with a novel domain from the hyperthermophile Thermotoga maritima

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Lei [Laboratory of Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Dreijenplein 10, 6703 HB Wageningen (Netherlands); Laboratory of Biophysical Chemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen (Netherlands); Levisson, Mark; Hendriks, Sjon; Akveld, Twan; Kengen, Servé W. M. [Laboratory of Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Dreijenplein 10, 6703 HB Wageningen (Netherlands); Dijkstra, Bauke W. [Laboratory of Biophysical Chemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen (Netherlands); Oost, John van der, E-mail: john.vanderoost@wur.nl [Laboratory of Microbiology, Department of Agrotechnology and Food Sciences, Wageningen University, Dreijenplein 10, 6703 HB Wageningen (Netherlands)

    2007-09-01

    A thermostable esterase (EstA) from Thermotoga maritima was cloned and purified. Crystals of EstA and its selenomethionine derivative were grown and diffract to beyond 2.6 Å resolution at 100 K using synchrotron radiation. A predicted esterase (EstA) with an unusual new domain from the hyperthermophilic bacterium Thermotoga maritima has been cloned and overexpressed in Escherichia coli. The purified protein was crystallized by the hanging-drop vapour-diffusion technique in the presence of lithium sulfate and polyethylene glycol 8000. Selenomethionine-substituted EstA crystals were obtained under the same conditions and three different-wavelength data sets were collected to 2.6 Å resolution. The crystal belongs to space group H32, with unit-cell parameters a = b = 130.2, c = 306.2 Å. There are two molecules in the asymmetric unit, with a V{sub M} of 2.9 Å{sup 3} Da{sup −1} and 58% solvent content.

  20. Crystallization and preliminary crystallographic studies of LipA, a secretory lipase/esterase from Xanthomonas oryzae pv. oryzae

    Energy Technology Data Exchange (ETDEWEB)

    Aparna, Gudlur; Chatterjee, Avradip; Jha, Gopaljee; Sonti, Ramesh V.; Sankaranarayanan, Rajan, E-mail: sankar@ccmb.res.in [Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad 500 007 (India)

    2007-08-01

    The crystallization and preliminary crystallographic studies of LipA, a lipase/esterase secreted by X. oryzae pv. oryzae during its infection of rice plants, are reported. Xanthomonas oryzae pv. oryzae is the causal agent of bacterial leaf blight, a serious disease of rice. Several enzymes that are secreted through the type II secretion system of this bacterium play an important role in the plant–microbe interaction, being important for virulence and also being able to induce potent host defence responses. One of these enzymes is a secretory lipase/esterase, LipA, which shows a very weak homology to other bacterial lipases and gives a positive tributyrin plate assay. In this study, LipA was purified from the culture supernatant of an overexpressing clone of X. oryzae pv. oryzae and two types of crystals belonging to space group C2 but with two different unit-cell parameters were obtained using the hanging-drop vapour-diffusion method. Type I crystals diffract to a maximum resolution of 1.89 Å and have unit-cell parameters a = 93.1, b = 62.3, c = 66.1 Å, β = 90.8°. Type II crystals have unit-cell parameters a = 103.6, b = 54.6, c = 66.3 Å, β = 92.6° and diffract to 1.86 Å. Solvent-content analysis shows one monomer in the asymmetric unit in both the crystal forms.

  1. Endophytic fungi producing of esterases: evaluation in vitro of the enzymatic activity using pH indicator

    Directory of Open Access Journals (Sweden)

    Helen Cristina Fávero Lisboa

    2013-09-01

    Full Text Available A sensitive and efficient colorimetric method was optimized for detection of esterase enzymes produced by endophytic fungi for development of High-Throughput Screening (HTS. The fungi were isolated and obtained previously from plant species of Cerrado and Atlantic Forest located in areas of environmental preservation in the State of Sao Paulo / Brazil, as part of the project "Chemical and biological prospecting endophytic fungi associated to plant species of Cerrado and Atlantic Forest". The compounds ethyl butyrate, ethyl acetate and methyl propionate were used as standards esters which were hydrolyzed by extracellular enzyme from endophytic fungi (EC. 3.1.1.1 -carboxylesterases for production of carboxylic acids. Thus, the reduction of the pH increases the protonated indicator concentration (bromothymol blue, changing the color of the reaction medium (from blue to yellow, that can be observed and measured by spectrophotometry at 616 nm. The methodology with acid-base indicator was performed on 13 microorganisms, aiming Periconia atropurpurea asapotential source of esterase for biotransformation of short chain esters. The results also evidenced that this methodology showed to be efficient, fast, cheap, having low consumption of reagents and easy development, and can be applied to screen carboxylic-ester hydrolases in a large number of microorganisms.

  2. Two distinct E3 ubiquitin ligases have complementary functions in the regulation of delta and serrate signaling in Drosophila.

    Directory of Open Access Journals (Sweden)

    Roland Le Borgne

    2005-04-01

    Full Text Available Signaling by the Notch ligands Delta (Dl and Serrate (Ser regulates a wide variety of essential cell-fate decisions during animal development. Two distinct E3 ubiquitin ligases, Neuralized (Neur and Mind bomb (Mib, have been shown to regulate Dl signaling in Drosophila melanogaster and Danio rerio, respectively. While the neur and mib genes are evolutionarily conserved, their respective roles in the context of a single organism have not yet been examined. We show here that the Drosophila mind bomb (D-mib gene regulates a subset of Notch signaling events, including wing margin specification, leg segmentation, and vein determination, that are distinct from those events requiring neur activity. D-mib also modulates lateral inhibition, a neur- and Dl-dependent signaling event, suggesting that D-mib regulates Dl signaling. During wing development, expression of D-mib in dorsal cells appears to be necessary and sufficient for wing margin specification, indicating that D-mib also regulates Ser signaling. Moreover, the activity of the D-mib gene is required for the endocytosis of Ser in wing imaginal disc cells. Finally, ectopic expression of neur in D-mib mutant larvae rescues the wing D-mib phenotype, indicating that Neur can compensate for the lack of D-mib activity. We conclude that D-mib and Neur are two structurally distinct proteins that have similar molecular activities but distinct developmental functions in Drosophila.

  3. The β-catenin E3 ubiquitin ligase SIAH-1 is regulated by CSN5/JAB1 in CRC cells.

    Science.gov (United States)

    Jumpertz, Sandra; Hennes, Thomas; Asare, Yaw; Vervoorts, Jörg; Bernhagen, Jürgen; Schütz, Anke K

    2014-09-01

    COP9 signalosome subunit 5 (CSN5) plays a decisive role in cellular processes such as cell cycle regulation and apoptosis via promoting protein degradation, gene transcription, and nuclear export. CSN5 regulates cullin-RING-E3 ligase (CRL) activity through its deNEDDylase function. It is overexpressed in several tumor entities, but its role in colorectal cancer (CRC) is poorly understood. Wnt/β-catenin signaling is aberrant in most CRC cells, resulting in increased levels of oncogenic β-catenin and thus tumor progression. Under physiological conditions, β-catenin levels are tightly regulated by continuous proteasomal degradation. We recently showed that knockdown of CSN5 in model and CRC cells results in decreased (phospho)-β-catenin levels. Reduced β-catenin levels were associated with an attenuated proliferation rate of different CRC cell types after CSN5 knockdown. The canonical Wnt pathway involves degradation of β-catenin by a β-TrCP1-containing E3 ligase, but is mostly non-functional in CRC cells. We thus hypothesized that alternative β-catenin degradation mediated by SIAH-1 (seven in absentia homolog-1), is responsible for the effect of CSN5 on β-catenin signaling in CRC cells. We found that SIAH-1 plays an essential role in β-catenin degradation in HCT116 CRC cells and that CSN5 affects β-catenin target gene expression in these cells. Of note, CSN5 affected SIAH-1 mRNA and SIAH-1 protein levels. Moreover, β-catenin and SIAH-1 form protein complexes with CSN5 in HCT116 cells. Lastly, we demonstrate that CSN5 promotes SIAH-1 degradation in HCT116 and SW480 cells and that this is associated with its deNEDDylase activity. In conclusion, we have identified a CSN5/β-catenin/SIAH-1 interaction network that might control β-catenin degradation in CRC cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  4. The interplay of descriptor-based computational analysis with pharmacophore modeling builds the basis for a novel classification scheme for feruloyl esterases

    DEFF Research Database (Denmark)

    Udatha, D.B.R.K. Gupta; Kouskoumvekaki, Irene; Olsson, Lisbeth

    2011-01-01

    One of the most intriguing groups of enzymes, the feruloyl esterases (FAEs), is ubiquitous in both simple and complex organisms. FAEs have gained importance in biofuel, medicine and food industries due to their capability of acting on a large range of substrates for cleaving ester bonds and synth...

  5. B-type esterases in the snail Xeropicta derbentina: An enzymological analysis to evaluate their use as biomarkers of pesticide exposure

    Energy Technology Data Exchange (ETDEWEB)

    Laguerre, Christel [Universite d' Avignon et des Pays de Vaucluse, UMR 406 UAPV/INRA, F-84914 Avignon (France); INRA, Laboratoire de Toxicologie Environnementale, UMR 406 UAPV/INRA, F-84914 Avignon (France); Sanchez-Hernandez, Juan C. [Laboratory of Ecotoxicology, Faculty of Environmental Science, University of Castilla-La Mancha, Avda. Carlos III s/n, 45071 Toledo (Spain); Koehler, Heinz R. [Animal Physiological Ecology, University of Tuebingen, Konrad-Adenauer-Strasse 20, D-72072 Tuebingen (Germany); Triebskorn, Rita [Animal Physiological Ecology, University of Tuebingen, Konrad-Adenauer-Strasse 20, D-72072 Tuebingen (Germany); Steinbeis-Transfer Center for Ecotoxicology and Ecophysiology, Blumenstrasse 13, D-72108 Rottenburg (Germany); Capowiez, Yvan [INRA, Unite PSH, F- 84914 Avignon (France); Rault, Magali [Universite d' Avignon et des Pays de Vaucluse, UMR 406 UAPV/INRA, F-84914 Avignon (France); INRA, Laboratoire de Toxicologie Environnementale, UMR 406 UAPV/INRA, F-84914 Avignon (France); Mazzia, Christophe [Universite d' Avignon et des Pays de Vaucluse, UMR 406 UAPV/INRA, F-84914 Avignon (France); INRA, Laboratoire de Toxicologie Environnementale, UMR 406 UAPV/INRA, F-84914 Avignon (France)], E-mail: mazzia@avignon.inra.fr

    2009-01-15

    The study was prompted to characterize the B-type esterase activities in the terrestrial snail Xeropicta derbentina and to evaluate its sensitivity to organophosphorus and carbamate pesticides. Specific cholinesterase and carboxylesterase activities were mainly obtained with acetylthiocholine (K{sub m} = 77.2 mM; V{sub max} = 38.2 mU/mg protein) and 1-naphthyl acetate (K{sub m} = 222 mM, V{sub max} = 1095 mU/mg protein) substrates, respectively. Acetylcholinesterase activity was concentration-dependently inhibited by chlorpyrifos-oxon, dichlorvos, carbaryl and carbofuran (IC50 = 1.35 x 10{sup -5}-3.80 x 10{sup -8} M). The organophosphate-inhibited acetylcholinesterase activity was reactivated in the presence of pyridine-2-aldoxime methochloride. Carboxylesterase activity was inhibited by organophosphorus insecticides (IC50 = 1.20 x 10{sup -5}-2.98 x 10{sup -8} M) but not by carbamates. B-esterase-specific differences in the inhibition by organophosphates and carbamates are discussed with respect to the buffering capacity of the carboxylesterase to reduce pesticide toxicity. These results suggest that B-type esterases in X. derbentina are suitable biomarkers of pesticide exposure and that this snail could be used as sentinel species in field monitoring of Mediterranean climate regions. - Characterization of the B-type esterases in the terrestrial snail Xeropicta derbentina in order to evaluate pesticide exposure.

  6. Plasma-derived human C1-esterase inhibitor does not prevent mechanical ventilation-induced pulmonary complement activation in a rat model of Streptococcus pneumoniae pneumonia

    NARCIS (Netherlands)

    de Beer, F. M.; Aslami, H.; Hoeksma, J.; van Mierlo, G.; Wouters, D.; Zeerleder, S.; Roelofs, J. J. T. H.; Juffermans, N. P.; Schultz, M. J.; Lagrand, W. K.

    2014-01-01

    Mechanical ventilation has the potential to cause lung injury, and the role of complement activation herein is uncertain. We hypothesized that inhibition of the complement cascade by administration of plasma-derived human C1-esterase inhibitor (C1-INH) prevents ventilation-induced pulmonary

  7. A small-volume bioassay for quantification of the esterase inhibiting potency of mixtures of organophosphate and carbamate insecticides in rainwater : development and optimization

    NARCIS (Netherlands)

    Hamers, T.; Molin, K.R.J.; Koeman, J.H.; Murk, A.J.

    2000-01-01

    The goal of this study was to develop a sensitive in vitro bioassay for quantification of the total esterase inhibiting potency of low concentrations of organophosphate and carbamate insecticides in relatively small rainwater samples. Purified acetylcholinesterase (AChE) from electric eel

  8. Esterase activity in the guinea pig thyroid under normal and pathological conditions (vitamin A deficiency) with special regard to cyst-like structures

    DEFF Research Database (Denmark)

    Kirkeby, S

    1977-01-01

    By use of different activators and inhibitors, TOCP(tri-o-cresyl phosphate), PCMB (parachloromercury benzoate), NiCl2, Pb(NO3)2, HgCl2, Hg(NO3)2, eserine and sodium taurocholate, it is shown that the esterase in the cyst cells and in group I cells of the guinea pig thyroid probably are A...

  9. Effects of piperonyl butoxide on the toxicity of the organophosphate temephos and the role of esterases in the insecticide resistance of Aedes aegypti

    Directory of Open Access Journals (Sweden)

    Boscolli Barbosa Pereira

    2014-10-01

    Full Text Available Introduction The effects of piperonyl butoxide (PBO on the toxicity of the organophosphate temephos (TE and the role of esterases in the resistance of Aedes aegypti to this insecticide were evaluated. Methods A. aegypti L4 larvae susceptible and resistant to TE were pre-treated with PBO solutions in acetone at concentrations of 0.125, 0.25, 0.5, 1, and 2% for 24h and subsequently exposed to a diagnostic concentration of 0.02mg/L aqueous TE solution. The esterase activity of the larvae extracts pre-treated with varying PBO concentrations and exposed to TE for three time periods was determined. Results At concentrations of 0.25, 0.5, 1, and 2%, PBO showed a significant synergistic effect with TE toxicity. High levels of esterase activity were associated with the survival of A. aegypti L4 larvae exposed to TE only. Conclusions The results of the biochemical assays suggest that PBO has a significant inhibitory effect on the total esterase activity in A. aegypti larvae.

  10. Histochemical studies on genetical control of hormonal enzyme inducibility in the mouse. I. Non-specific esterase activity and regional histology of the epididymis

    DEFF Research Database (Denmark)

    Blecher, S R; Kirkeby, S

    1978-01-01

    As a base line for future cell genetical studies the authors record the distribution of non-specific esterase reaction in the various histologically distinguishable cell types of the mouse epididymis. The findings are correlated with previous descriptions of the lobar structure of the organ...

  11. Connexin31.1 deficiency in the mouse impairs object memory and modulates open-field exploration, acetylcholine esterase levels in the striatum, and cAMP response element-binding protein levels in the striatum and piriform cortex.

    Science.gov (United States)

    Dere, E; Zheng-Fischhöfer, Q; Viggiano, D; Gironi Carnevale, U A; Ruocco, L A; Zlomuzica, A; Schnichels, M; Willecke, K; Huston, J P; Sadile, A G

    2008-05-02

    Neuronal gap junctions in the brain, providing intercellular electrotonic signal transfer, have been implicated in physiological and behavioral correlates of learning and memory. In connexin31.1 (Cx31.1) knockout (KO) mice the coding region of the Cx31.1 gene was replaced by a LacZ reporter gene. We investigated the impact of Cx31.1 deficiency on open-field exploration, the behavioral response to an odor, non-selective attention, learning and memory performance, and the levels of memory-related proteins in the hippocampus, striatum and the piriform cortex. In terms of behavior, the deletion of the Cx31.1 coding DNA in the mouse led to increased exploratory behaviors in a novel environment, and impaired one-trial object recognition at all delays tested. Despite strong Cx31.1 expression in the peripheral and central olfactory system, Cx31.1 KO mice exhibited normal behavioral responses to an odor. We found increased levels of acetylcholine esterase (AChE) and cAMP response element-binding protein (CREB) in the striatum of Cx31.1 KO mice. In the piriform cortex the Cx31.1 KO mice had an increased heterogeneity of CREB expression among neurons. In conclusion, gap-junctions featuring the Cx31.1 protein might be involved in open-field exploration as well as object memory and modulate levels of AChE and CREB in the striatum and piriform cortex.

  12. Case study of the interdisciplinary integration in an IST-E3 project

    DEFF Research Database (Denmark)

    Jørgensen, Ulrik

    2002-01-01

    Case study of a specific IST-E3 project funded by the EU commissions 5th framework program. The case study highlights the difficulties in integrating different disciplinary approaches and suggests that a more openended research strategy should be applied by the commission.......Case study of a specific IST-E3 project funded by the EU commissions 5th framework program. The case study highlights the difficulties in integrating different disciplinary approaches and suggests that a more openended research strategy should be applied by the commission....

  13. Bioinformatics analysis identifies several intrinsically disordered human E3 ubiquitin-protein ligases

    Directory of Open Access Journals (Sweden)

    Wouter Boomsma

    2016-02-01

    Full Text Available The ubiquitin-proteasome system targets misfolded proteins for degradation. Since the accumulation of such proteins is potentially harmful for the cell, their prompt removal is important. E3 ubiquitin-protein ligases mediate substrate ubiquitination by bringing together the substrate with an E2 ubiquitin-conjugating enzyme, which transfers ubiquitin to the substrate. For misfolded proteins, substrate recognition is generally delegated to molecular chaperones that subsequently interact with specific E3 ligases. An important exception is San1, a yeast E3 ligase. San1 harbors extensive regions of intrinsic disorder, which provide both conformational flexibility and sites for direct recognition of misfolded targets of vastly different conformations. So far, no mammalian ortholog of San1 is known, nor is it clear whether other E3 ligases utilize disordered regions for substrate recognition. Here, we conduct a bioinformatics analysis to examine >600 human and S. cerevisiae E3 ligases to identify enzymes that are similar to San1 in terms of function and/or mechanism of substrate recognition. An initial sequence-based database search was found to detect candidates primarily based on the homology of their ordered regions, and did not capture the unique disorder patterns that encode the functional mechanism of San1. However, by searching specifically for key features of the San1 sequence, such as long regions of intrinsic disorder embedded with short stretches predicted to be suitable for substrate interaction, we identified several E3 ligases with these characteristics. Our initial analysis revealed that another remarkable trait of San1 is shared with several candidate E3 ligases: long stretches of complete lysine suppression, which in San1 limits auto-ubiquitination. We encode these characteristic features into a San1 similarity-score, and present a set of proteins that are plausible candidates as San1 counterparts in humans. In conclusion, our work

  14. Monitoring of high-risk pregnancies using E3 and HPL

    Energy Technology Data Exchange (ETDEWEB)

    Bieglmayer, Ch; Spona, J

    1981-01-01

    Routine determinations of HPL and E3 together with other clinical and laboratory checks offer usable information on the state of pregnancy and assist the doctor in taking decisions on the use of therapeutical measures. The anamnesis, the presence of placental insufficiency of EPH-gestosis are important indications for monitoring the pregnancy using HPL and E3. A commercial RIA apparatus (Centria 2) allowing the determination of HPL in a mere hour was compared with our established laboratory method. In 140 determinations, the correlation was r=0.75. Considering its capacity and method, the instrument seems to be more suitable for a small specialized laboratory.

  15. Interactive toxicity of chlorpyrifos and parathion in neonatal rats: Role of esterases in exposure sequence-dependent toxicity

    International Nuclear Information System (INIS)

    Kacham, R.; Karanth, S.; Baireddy, P.; Liu, J.; Pope, C.

    2006-01-01

    We previously reported that sequence of exposure to chlorpyrifos and parathion in adult rats can markedly influence toxic outcome. In the present study, we evaluated the interactive toxicity of chlorpyrifos (8 mg/kg, po) and parathion (0.5 mg/kg, po) in neonatal (7 days old) rats. Rats were exposed to the insecticides either concurrently or sequentially (separated by 4 h) and sacrificed at 4, 8, and 24 h after the first exposure for biochemical measurements (cholinesterase activity in brain, plasma, and diaphragm and carboxylesterase activity in plasma and liver). The concurrently-exposed group showed more cumulative lethality (15/24) than either of the sequential dosing groups. With sequential dosing, rats treated initially with chlorpyrifos prior to parathion (C/P) exhibited higher lethality (7/23) compared to those treated with parathion before chlorpyrifos (P/C; 1/24). At 8 h after initial dosing, brain cholinesterase inhibition was significantly greater in the C/P group (59%) compared to the P/C group (28%). Diaphragm and plasma cholinesterase activity also followed a relatively similar pattern of inhibition. Carboxylesterase inhibition in plasma and liver was relatively similar among the treatment groups across time-points. Similar sequence-dependent differences in brain cholinesterase inhibition were also noted with lower binary exposures to chlorpyrifos (2 mg/kg) and parathion (0.35 mg/kg). In vitro and ex vivo studies compared relative oxon detoxification of carboxylesterases (calcium-insensitive) and A-esterases (calcium-sensitive) in liver homogenates from untreated and insecticide pretreated rats. Using tissues from untreated rats, carboxylesterases detoxified both chlorpyrifos oxon and paraoxon, while A-esterases only detoxified chlorpyrifos oxon. With parathion pretreatment, A-esterases still detoxified chlorpyrifos oxon while liver from chlorpyrifos pretreated rats had little apparent effect on paraoxon. We conclude that while neonatal rats are less

  16. Esterase polymorphism in remanant populations of Aspidosperma polyneuron Müll.Arg. (Apocynaceae Polimorfismo de esterases em populações remanescentes de Aspidosperma polyneuron Müll.Arg. (Apocynaceae

    Directory of Open Access Journals (Sweden)

    Vanda Marilza de Carvalho

    2004-10-01

    Full Text Available The population genetic structure of the endangered tree species Aspidosperma polyneuron Mull.Arg. (Apocynaceae was reported based on analysis of esterase polymorphism in two remanant populations. Allelic variation was detected at three isoesterase loci (Est-3, Est-9, and Est-10. The proportion of polymorphic loci for both populations was 30% and deviation from Hardy-Weinberg equilibrium was observed for the Est-3 locus observed in the northern population. Segregation distortion and the lower level of observed and expected heterozygosity in this population were attributed to founder genotype. The high genetic identity values for northern and northwestern populations are in accordance with the low levels of interpopulation genetic divergence demonstrated by the F(ST (0.03 value. The F(IS value (0.23 indicated moderate levels of inbreeding. A. polyneuron can be indicated as an example of endangered species suggesting high genetic variation in contrast to the low genetic variation reported for endangered species. The esterase isozymes may be a good genetic marker for studies of natural A. polyneuron populations.A análise do polimorfismo de isozimas esterases foi usada para reportar a estrutura genética de duas populações remanecentes da espécie de árvore em extinção Aspidosperma polyneuron Müll.Arg. (Apocynaceae. Variação alélica foi detectada em três locos de isoesterases (Est-3, Est-9, e Est-10. A proporção de locos polimórficos de ambas as populações foi de 30%, sendo observado um desvio do equilíbrio de Hardy-Weinberg no loco Est-3 na população da região norte do Estado do Paraná. Uma distorção na segregação e um mais baixo nível de heterozigosidade observada e esperada nesta população foram atribuídos ao efeito do genótipo fundador. Os valores altos de identidade genética das populações do norte e noroeste do Estado estão de acordo com o baixo nível de divergência genética interpopulacional demonstrado

  17. Detection of carboxylesterase and esterase activity in culturable gut bacterial flora isolated from diamondback moth, Plutella xylostella (Linnaeus), from India and its possible role in indoxacarb degradation.

    Science.gov (United States)

    Ramya, Shanivarsanthe Leelesh; Venkatesan, Thiruvengadam; Srinivasa Murthy, Kottilingam; Jalali, Sushil Kumar; Verghese, Abraham

    2016-01-01

    Diamondback moth (DBM), Plutella xylostella (Linnaeus), is a notorious pest of brassica crops worldwide and is resistant to all groups of insecticides. The insect system harbors diverse groups of microbiota, which in turn helps in enzymatic degradation of xenobiotic-like insecticides. The present study aimed to determine the diversity of gut microflora in DBM, quantify esterase activity and elucidate their possible role in degradation of indoxacarb. We screened 11 geographic populations of DBM in India and analyzed them for bacterial diversity. The culturable gut bacterial flora underwent molecular characterization with 16S rRNA. We obtained 25 bacterial isolates from larvae (n=13) and adults (n=12) of DBM. In larval gut isolates, gammaproteobacteria was the most abundant (76%), followed by bacilli (15.4%). Molecular characterization placed adult gut bacterial strains into three major classes based on abundance: gammaproteobacteria (66%), bacilli (16.7%) and flavobacteria (16.7%). Esterase activity from 19 gut bacterial isolates ranged from 0.072 to 2.32μmol/min/mg protein. Esterase bands were observed in 15 bacterial strains and the banding pattern differed in Bacillus cereus - KC985225 and Pantoea agglomerans - KC985229. The bands were characterized as carboxylesterase with profenofos used as an inhibitor. Minimal media study showed that B. cereus degraded indoxacarb up to 20%, so it could use indoxacarb for metabolism and growth. Furthermore, esterase activity was greater with minimal media than control media: 1.87 versus 0.26μmol/min/mg protein. Apart from the insect esterases, bacterial carboxylesterase may aid in the degradation of insecticides in DBM. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  18. Detection of carboxylesterase and esterase activity in culturable gut bacterial flora isolated from diamondback moth, Plutella xylostella (Linnaeus, from India and its possible role in indoxacarb degradation

    Directory of Open Access Journals (Sweden)

    Shanivarsanthe Leelesh Ramya

    2016-06-01

    Full Text Available Abstract Diamondback moth (DBM, Plutella xylostella (Linnaeus, is a notorious pest of brassica crops worldwide and is resistant to all groups of insecticides. The insect system harbors diverse groups of microbiota, which in turn helps in enzymatic degradation of xenobiotic-like insecticides. The present study aimed to determine the diversity of gut microflora in DBM, quantify esterase activity and elucidate their possible role in degradation of indoxacarb. We screened 11 geographic populations of DBM in India and analyzed them for bacterial diversity. The culturable gut bacterial flora underwent molecular characterization with 16S rRNA. We obtained 25 bacterial isolates from larvae (n = 13 and adults (n = 12 of DBM. In larval gut isolates, gammaproteobacteria was the most abundant (76%, followed by bacilli (15.4%. Molecular characterization placed adult gut bacterial strains into three major classes based on abundance: gammaproteobacteria (66%, bacilli (16.7% and flavobacteria (16.7%. Esterase activity from 19 gut bacterial isolates ranged from 0.072 to 2.32 µmol/min/mg protein. Esterase bands were observed in 15 bacterial strains and the banding pattern differed in Bacillus cereus – KC985225 and Pantoea agglomerans – KC985229. The bands were characterized as carboxylesterase with profenofos used as an inhibitor. Minimal media study showed that B. cereus degraded indoxacarb up to 20%, so it could use indoxacarb for metabolism and growth. Furthermore, esterase activity was greater with minimal media than control media: 1.87 versus 0.26 µmol/min/mg protein. Apart from the insect esterases, bacterial carboxylesterase may aid in the degradation of insecticides in DBM.

  19. TMEM129 is a Derlin-1 associated ERAD E3 ligase essential for virus-induced degradation of MHC-I.

    Science.gov (United States)

    van den Boomen, Dick J H; Timms, Richard T; Grice, Guinevere L; Stagg, Helen R; Skødt, Karsten; Dougan, Gordon; Nathan, James A; Lehner, Paul J

    2014-08-05

    The US11 gene product of human cytomegalovirus promotes viral immune evasion by hijacking the endoplasmic reticulum (ER)-associated degradation (ERAD) pathway. US11 initiates dislocation of newly translocated MHC I from the ER to the cytosol for proteasome-mediated degradation. Despite the critical role for ubiquitin in this degradation pathway, the responsible E3 ligase is unknown. In a forward genetic screen for host ERAD components hijacked by US11 in near-haploid KBM7 cells, we identified TMEM129, an uncharacterized polytopic membrane protein. TMEM129 is essential and rate-limiting for US11-mediated MHC-I degradation and acts as a novel ER resident E3 ubiquitin ligase. TMEM129 contains an unusual cysteine-only RING with intrinsic E3 ligase activity and is recruited to US11 via Derlin-1. Together with its E2 conjugase Ube2J2, TMEM129 is responsible for the ubiquitination, dislocation, and subsequent degradation of US11-associated MHC-I. US11 engages two degradation pathways: a Derlin-1/TMEM129-dependent pathway required for MHC-I degradation and a SEL1L/HRD1-dependent pathway required for "free" US11 degradation. Our data show that TMEM129 is a novel ERAD E3 ligase and the central component of a novel mammalian ERAD complex.

  20. LOSS OF JAK2 REGULATION VIA VHL-SOCS1 E3 UBIQUITIN HETEROCOMPLEX UNDERLIES CHUVASH POLYCYTHEMIA

    Science.gov (United States)

    Russell, Ryan C.; Sufan, Roxana I.; Zhou, Bing; Heir, Pardeep; Bunda, Severa; Sybingco, Stephanie S.; Greer, Samantha N.; Roche, Olga; Heathcote, Samuel A.; Chow, Vinca W.K.; Boba, Lukasz M.; Richmond, Terri D.; Hickey, Michele M.; Barber, Dwayne L.; Cheresh, David A.; Simon, M. Celeste; Irwin, Meredith S.; Kim, William Y.; Ohh, Michael

    2011-01-01

    SUMMARY Chuvash polycythemia (CP) is a rare congenital form of polycythemia caused by homozygous R200W and H191D mutations in the von Hippel-Lindau (VHL) gene whose gene product is the principal negative regulator of hypoxia-inducible factor. However, the molecular mechanisms underlying some of the hallmark features of CP such as hypersensitivity to erythropoietin are unclear. Here, we show that VHL directly binds suppressor of cytokine signalling 1 (SOCS1) to form a heterodimeric E3 ligase that targets phosphorylated (p)JAK2 for ubiquitin-mediated destruction. In contrast, CP-associated VHL mutants have altered affinity for SOCS1 and fail to engage and degrade pJAK2. Systemic administration of a highly selective JAK2 inhibitor, TG101209, reverses the disease phenotype in vhlR200W/R200W knock-in mice, a model that faithfully recapitulates human CP. These results reveal VHL as a SOCS1-cooperative negative regulator of JAK2 and provide compelling biochemical and preclinical evidence for JAK2- targeted therapy in CP patients. PMID:21685897

  1. Bioinformatics analysis identifies several intrinsically disordered human E3 ubiquitin-protein ligases

    DEFF Research Database (Denmark)

    Boomsma, Wouter Krogh; Nielsen, Sofie Vincents; Lindorff-Larsen, Kresten

    2016-01-01

    conduct a bioinformatics analysis to examine >600 human and S. cerevisiae E3 ligases to identify enzymes that are similar to San1 in terms of function and/or mechanism of substrate recognition. An initial sequence-based database search was found to detect candidates primarily based on the homology...

  2. NMR characterization of foldedness for the production of E3 RING domains

    NARCIS (Netherlands)

    Huang, A.; de Jong, R.N.; Folkers, G.E.; Boelens, R.

    2010-01-01

    We summarize the use of NMR spectroscopy in the production and the screening of stability and foldedness of protein domains, and apply it to the RING domains of E3 ubiquitin-ligases. RING domains are involved in specific interactions with E2 ubiquitin-conjugating enzymes and thus play an essential

  3. Ophthalmic epidemiology in Europe : the "European Eye Epidemiology" (E3) consortium

    NARCIS (Netherlands)

    Delcourt, Cecile; Korobelnik, Jean-Francois; Buitendijk, Gabrielle H. S.; Foster, Paul J.; Hammond, Christopher J.; Piermarocchi, Stefano; Peto, Tunde; Jansonius, Nomdo; Mirshahi, Alireza; Hogg, Ruth E.; Bretillon, Lionel; Topouzis, Fotis; Deak, Gabor; Grauslund, Jakob; Broe, Rebecca; Souied, Eric H.; Creuzot-Garcher, Catherine; Sahel, Jose; Daien, Vincent; Lehtimaki, Terho; Hense, Hans-Werner; Prokofyeva, Elena; Oexle, Konrad; Rahi, Jugnoo S.; Cumberland, Phillippa M.; Schmitz-Valckenberg, Steffen; Fauser, Sascha; Bertelsen, Geir; Hoyng, Carel; Bergen, Arthur; Silva, Rufino; Wolf, Sebastian; Lotery, Andrew; Chakravarthy, Usha; Fletcher, Astrid; Klaver, Caroline C. W.

    The European Eye Epidemiology (E3) consortium is a recently formed consortium of 29 groups from 12 European countries. It already comprises 21 population-based studies and 20 other studies (case-control, cases only, randomized trials), providing ophthalmological data on approximately 170,000

  4. Dual RING E3 Architectures Regulate Multiubiquitination and Ubiquitin Chain Elongation by APC/C.

    Science.gov (United States)

    Brown, Nicholas G; VanderLinden, Ryan; Watson, Edmond R; Weissmann, Florian; Ordureau, Alban; Wu, Kuen-Phon; Zhang, Wei; Yu, Shanshan; Mercredi, Peter Y; Harrison, Joseph S; Davidson, Iain F; Qiao, Renping; Lu, Ying; Dube, Prakash; Brunner, Michael R; Grace, Christy R R; Miller, Darcie J; Haselbach, David; Jarvis, Marc A; Yamaguchi, Masaya; Yanishevski, David; Petzold, Georg; Sidhu, Sachdev S; Kuhlman, Brian; Kirschner, Marc W; Harper, J Wade; Peters, Jan-Michael; Stark, Holger; Schulman, Brenda A

    2016-06-02

    Protein ubiquitination involves E1, E2, and E3 trienzyme cascades. E2 and RING E3 enzymes often collaborate to first prime a substrate with a single ubiquitin (UB) and then achieve different forms of polyubiquitination: multiubiquitination of several sites and elongation of linkage-specific UB chains. Here, cryo-EM and biochemistry show that the human E3 anaphase-promoting complex/cyclosome (APC/C) and its two partner E2s, UBE2C (aka UBCH10) and UBE2S, adopt specialized catalytic architectures for these two distinct forms of polyubiquitination. The APC/C RING constrains UBE2C proximal to a substrate and simultaneously binds a substrate-linked UB to drive processive multiubiquitination. Alternatively, during UB chain elongation, the RING does not bind UBE2S but rather lures an evolving substrate-linked UB to UBE2S positioned through a cullin interaction to generate a Lys11-linked chain. Our findings define mechanisms of APC/C regulation, and establish principles by which specialized E3-E2-substrate-UB architectures control different forms of polyubiquitination. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Characterizations of Space Curves According to Bishop Darboux Vector in Euclidean 3-Space E3

    OpenAIRE

    Huseyin KOCAYIGIT; Ali OZDEMIR

    2014-01-01

    In this paper, we obtained some characterizations of space curves according to Bihop frame in Euclidean 3-space E3 by using Laplacian operator and Levi-Civita connection. Furthermore, we gave the general differential equations which characterize the space curves according to the Bishop Darboux vector and the normal Bishop Darboux vector.

  6. SH groups in the alpha-naphthyl acetate esterase in the thyroid of the guinea-pig. A histochemical study

    DEFF Research Database (Denmark)

    Kirkeby, S

    1976-01-01

    ] are added to the incubation media. Thus the inhibition is by far the greatest in group I cells, which also show the greatest activity after incubation in conventional media, when long fixation and storage times are used. In all cases the inhibiting effect was complete or almost completely reversed......The alpha-naphthyl acetate esterase in both group I and group II thyroid cells is shown to contain SH groups since there is a decline in activity in both cell groups when certain sulfhydryl reagents [DTNB; 5,5'-Dithiobis-(2-nitrobenzoic acid)-AgNO3-Mersalyl-PCMB (parachloro mercuribenzoate) + urea...... if cysteine was added to the incubation media in equivalent concentrations to the SH blocker. There were great differences among the sulfhydryl reagents used in their ability to bring about enzyme inhibition. The alkylating agents NEM (N-ethylmaleimide) and iodoacetamide had no or little effect while PCMB...

  7. Effect of temperature and high pressure on the activity and mode of action of fungal pectin methyl esterase.

    Science.gov (United States)

    Duvetter, Thomas; Fraeye, Ilse; Sila, Daniel N; Verlent, Isabel; Smout, Chantal; Clynen, Elke; Schoofs, Liliane; Schols, Henk; Hendrickx, Marc; Van Loey, Ann

    2006-01-01

    Pectin was de-esterified with purified recombinant Aspergillus aculeatus pectin methyl esterase (PME) during isothermal-isobaric treatments. By measuring the release of methanol as a function of treatment time, the rate of enzymatic pectin conversion was determined. Elevated temperature and pressure were found to stimulate PME activity. The highest rate of PME-catalyzed pectin de-esterification was obtained when combining pressures in the range 200-300 MPa with temperatures in the range 50-55 degrees C. The mode of pectin de-esterification was investigated by characterizing the pectin reaction products by enzymatic fingerprinting. No significant effect of increasing pressure (300 MPa) and/or temperature (50 degrees C) on the mode of pectin conversion was detected.

  8. Pectin methyl esterase and natural microflora of fresh mixed orange and carrot juice treated with pulsed electric fields.

    Science.gov (United States)

    Rodrigo, D; Barbosa-Cánovas, G V; Martínez, A; Rodrigo, M

    2003-12-01

    The effects of pulsed electric fields (PEFs) on pectin methyl esterase (PME), molds and yeast, and total flora in fresh (nonpasteurized) mixed orange and carrot juice were studied. The PEF effect was more extensive when juices with high levels of initial PME activity were subjected to treatment and when PEF treatment (at 25 kV/cm for 340 micros) was combined with a moderate temperature (63 degrees C), with the maximum level of PME inactivation being 81.4%. These conditions produced 3.7 decimal reductions in molds and yeast and 2.4 decimal reductions in total flora. Experimental inactivation data for PME, molds and yeast, and total flora were fitted to Bigelow, Hülsheger, and Weibull inactivation models by nonlinear regression. The best fit (lowest mean square error) was obtained with the Weibull model.

  9. Xylanase and feruloyl esterase from actinomycetes cultures could enhance sugarcane bagasse hydrolysis in the production of fermentable sugars.

    Science.gov (United States)

    Rahmani, Nanik; Kahar, Prihardi; Lisdiyanti, Puspita; Hermiati, Euis; Lee, Jaemin; Yopi; Prasetya, Bambang; Ogino, Chiaki; Kondo, Akihiko

    2018-02-23

    The addition of enzymes that are capable of degrading hemicellulose has a potential to reduce the need for commercial enzymes during biomass hydrolysis in the production of fermentable sugars. In this study, a high xylanase producing actinomycete strain (Kitasatospora sp. ID06-480) and the first ethyl ferulate producing actinomycete strain (Nonomuraea sp. ID06-094) were selected from 797 rare actinomycetes, respectively, which were isolated in Indonesia. The addition (30%, v/v) of a crude enzyme supernatant from the selected strains in sugarcane bagasse hydrolysis with low-level loading (1 FPU/g-biomass) of Cellic® CTec2 enhanced both the released amount of glucose and reducing sugars. When the reaction with Ctec2 was combined with crude enzymes containing either xylanase or feruloyl esterase, high conversion yield of glucose from cellulose at 60.5% could be achieved after 72 h-saccharification.

  10. Immobilization of cholesterol esterase and cholesterol oxidase onto sol-gel films for application to cholesterol biosensor

    International Nuclear Information System (INIS)

    Singh, Suman; Singhal, Rahul; Malhotra, B.D.

    2007-01-01

    Cholesterol oxidase (ChOx) and cholesterol esterase (ChEt) have been covalently immobilized onto tetraethylorthosilicate (TEOS) sol-gel films. The tetraethylorthosilicate sol-gel/ChEt/ChOx enzyme films thus prepared have been characterized using scanning electron microscopic (SEM), UV-vis spectroscopic, Fourier-transform-infrared (FTIR) spectroscopic and amperometric techniques, respectively. The results of photometric measurements carried out on tetraethylorthosilicate sol-gel/ChEt/ChOx reveal thermal stability up to 55 deg. C, response time as 180 s, linearity up to 780 mg dL -1 (12 mM), shelf life of 1 month, detection limit of 12 mg dL -1 and sensitivity as 5.4 x 10 -5 Abs. mg -1 dL -1

  11. Immobilization of cholesterol esterase and cholesterol oxidase onto sol-gel films for application to cholesterol biosensor

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Suman [Central Mechanical Engineering Research Institute, G. Avenue, Durgapur 713209, West Bengal (India); Singhal, Rahul [Biomolecular Electronics and Conducting Polymer Research Group, National Physical Laboratory, Dr. K.S. Krishnan Marg, New Delhi 110012 (India); Malhotra, B.D. [Biomolecular Electronics and Conducting Polymer Research Group, National Physical Laboratory, Dr. K.S. Krishnan Marg, New Delhi 110012 (India)]. E-mail: bansi.malhotra@gmail.com

    2007-01-23

    Cholesterol oxidase (ChOx) and cholesterol esterase (ChEt) have been covalently immobilized onto tetraethylorthosilicate (TEOS) sol-gel films. The tetraethylorthosilicate sol-gel/ChEt/ChOx enzyme films thus prepared have been characterized using scanning electron microscopic (SEM), UV-vis spectroscopic, Fourier-transform-infrared (FTIR) spectroscopic and amperometric techniques, respectively. The results of photometric measurements carried out on tetraethylorthosilicate sol-gel/ChEt/ChOx reveal thermal stability up to 55 deg. C, response time as 180 s, linearity up to 780 mg dL{sup -1} (12 mM), shelf life of 1 month, detection limit of 12 mg dL{sup -1} and sensitivity as 5.4 x 10{sup -5} Abs. mg{sup -1} dL{sup -1}.

  12. Pathfinding the Flight Advanced Stirling Convertor Design with the ASC-E3

    Science.gov (United States)

    Wong, Wayne A.; Wilson, Kyle; Smith, Eddie; Collins, Josh

    2012-01-01

    The Advanced Stirling Convertor (ASC) was initially developed by Sunpower, Inc. under contract to NASA Glenn Research Center (GRC) as a technology development project. The ASC technology fulfills NASA's need for high efficiency power convertors for future Radioisotope Power Systems (RPS). Early successful technology demonstrations between 2003 to 2005 eventually led to the expansion of the project including the decision in 2006 to use the ASC technology on the Advanced Stirling Radioisotope Generator (ASRG). Sunpower has delivered 22 ASC convertors of progressively mature designs to date to GRC. Currently, Sunpower with support from GRC, Lockheed Martin Space System Company (LMSSC), and the Department of Energy (DOE) is developing the flight ASC-F in parallel with the ASC-E3 pathfinders. Sunpower will deliver four pairs of ASC-E3 convertors to GRC which will be used for extended operation reliability assessment, independent validation and verification testing, system interaction tests, and to support LMSSC controller verification. The ASC-E3 and -F convertors are being built to the same design and processing documentation and the same product specification. The initial two pairs of ASC-E3 are built before the flight units and will validate design and processing changes prior to implementation on the ASC-F flight convertors. This paper provides a summary on development of the ASC technology and the status of the ASC-E3 build and how they serve the vital pathfinder role ahead of the flight build for ASRG. The ASRG is part of two of the three candidate missions being considered for selection for the Discovery 12 mission.

  13. A Thermally Stable Form of Bacterial Cocaine Esterase: A Potential Therapeutic Agent for Treatment of Cocaine Abuse

    Energy Technology Data Exchange (ETDEWEB)

    Brim, Remy L.; Nance, Mark R.; Youngstrom, Daniel W.; Narasimhan, Diwahar; Zhan, Chang-Guo; Tesmer, John J.G.; Sunahara, Roger K.; Woods, James H. (Michigan); (Michigan-Med); (Kentucky)

    2010-09-03

    Rhodococcal cocaine esterase (CocE) is an attractive potential treatment for both cocaine overdose and cocaine addiction. CocE directly degrades cocaine into inactive products, whereas traditional small-molecule approaches require blockade of the inhibitory action of cocaine on a diverse array of monoamine transporters and ion channels. The usefulness of wild-type (wt) cocaine esterase is hampered by its inactivation at 37 C. Herein, we characterize the most thermostable form of this enzyme to date, CocE-L169K/G173Q. In vitro kinetic analyses reveal that CocE-L169K/G173Q displays a half-life of 2.9 days at 37 C, which represents a 340-fold improvement over wt and is 15-fold greater than previously reported mutants. Crystallographic analyses of CocE-L169K/G173Q, determined at 1.6-{angstrom} resolution, suggest that stabilization involves enhanced domain-domain interactions involving van der Waals interactions and hydrogen bonding. In vivo rodent studies reveal that intravenous pretreatment with CocE-L169K/G173Q in mice provides protection from cocaine-induced lethality for longer time periods before cocaine administration than wt CocE. Furthermore, intravenous administration (pretreatment) of CocE-L169K/G173Q prevents self-administration of cocaine in a time-dependent manner. Termination of the in vivo effects of CoCE seems to be dependent on, but not proportional to, its clearance from plasma as its half-life is approximately 2.3 h and similar to that of wt CocE (2.2 h). Taken together these data suggest that CocE-L169K/G173Q possesses many of the properties of a biological therapeutic for treating cocaine abuse but requires additional development to improve its serum half-life.

  14. Down-Regulation of Neuropathy Target Esterase in Preeclampsia Placenta Inhibits Human Trophoblast Cell Invasion via Modulating MMP-9 Levels

    Directory of Open Access Journals (Sweden)

    Ting Zhong

    2018-02-01

    Full Text Available Background/Aims: Neuropathy target esterase (NTE, also known as neurotoxic esterase is proven to deacylate phosphatidylcholine (PC to glycerophosphocholine as a phospholipase B. Recently; studies showed that artificial phosphatidylserine/PC microvesicles can induce preeclampsia (PE-like changes in pregnant mice. However, it is unclear whether NTE plays a key role in the pathology of PE, a pregnancy-related disease, which was characterized by deficient trophoblast invasion and reduced trophoblast-mediated remodeling of spiral arteries. The aim of this study was to investigate the expression pattern of NTE in the placenta from women with PE and normal pregnancy, and the molecular mechanism of NTE involved in the development of PE. Methods: NTE expression levels in placentas from 20 pregnant women with PE and 20 healthy pregnant women were detected using quantitative PCR and immunohistochemistry staining. The effect of NTE on trophoblast migration and invasion and the underlying mechanisms were examined in HTR-8/SVneo cell lines by transfection method. Results: NTE mRNA and protein expression levels were significantly decreased in preeclamptic placentas than normal control. Over-expression of NTE in HTR-8/SVneo cells significantly promoted trophoblast cells migration and invasion and was associated with increased MMP-9 levels. Conversely, shRNA-mediated down-regulation of NTE markedly inhibited the cell migration and invasion. In addition, silencing NTE reduced the MMP-9 activity and phosphorylated Erk1/2 and AKT levels. Conclusions: Our results suggest that the decreased NTE may contribute to the development of PE through impairing trophoblast invasion by down-regulating MMP-9 via the Erk1/2 and AKT signaling pathway.

  15. Bacillus sp. JR3 esterase LipJ: A new mesophilic enzyme showing traces of a thermophilic past.

    Directory of Open Access Journals (Sweden)

    Judit Ribera

    Full Text Available A search for extremophile enzymes from ancient volcanic soils in El Hierro Island (Canary Islands, Spain allowed isolation of a microbial sporulated strain collection from which several enzymatic activities were tested. Isolates were obtained after sample cultivation under several conditions of nutrient contents and temperature. Among the bacterial isolates, supernatants from the strain designated JR3 displayed high esterase activity at temperatures ranging from 30 to 100°C, suggesting the presence of at least a hyper-thermophilic extracellular lipase. Sequence alignment of known thermophilic lipases allowed design of degenerated consensus primers for amplification and cloning of the corresponding lipase, named LipJ. However, the cloned enzyme displayed maximum activity at 30°C and pH 7, showing a different profile from that observed in supernatants of the parental strain. Sequence analysis of the cloned protein showed a pentapeptide motif -GHSMG- distinct from that of thermophilic lipases, and much closer to that of esterases. Nevertheless, the 3D structural model of LipJ displayed the same folding as that of thermophilic lipases, suggesting a common evolutionary origin. A phylogenetic study confirmed this possibility, positioning LipJ as a new member of the thermophilic family of bacterial lipases I.5. However, LipJ clusters in a clade close but separated from that of Geobacillus sp. thermophilic lipases. Comprehensive analysis of the cloned enzyme suggests a common origin of LipJ and other bacterial thermophilic lipases, and highlights the most probable divergent evolutionary pathway followed by LipJ, which during the harsh past times would have probably been a thermophilic enzyme, having lost these properties when the environment changed to more benign conditions.

  16. Bacillus sp. JR3 esterase LipJ: A new mesophilic enzyme showing traces of a thermophilic past.

    Science.gov (United States)

    Ribera, Judit; Estupiñán, Mónica; Fuentes, Alba; Fillat, Amanda; Martínez, Josefina; Diaz, Pilar

    2017-01-01

    A search for extremophile enzymes from ancient volcanic soils in El Hierro Island (Canary Islands, Spain) allowed isolation of a microbial sporulated strain collection from which several enzymatic activities were tested. Isolates were obtained after sample cultivation under several conditions of nutrient contents and temperature. Among the bacterial isolates, supernatants from the strain designated JR3 displayed high esterase activity at temperatures ranging from 30 to 100°C, suggesting the presence of at least a hyper-thermophilic extracellular lipase. Sequence alignment of known thermophilic lipases allowed design of degenerated consensus primers for amplification and cloning of the corresponding lipase, named LipJ. However, the cloned enzyme displayed maximum activity at 30°C and pH 7, showing a different profile from that observed in supernatants of the parental strain. Sequence analysis of the cloned protein showed a pentapeptide motif -GHSMG- distinct from that of thermophilic lipases, and much closer to that of esterases. Nevertheless, the 3D structural model of LipJ displayed the same folding as that of thermophilic lipases, suggesting a common evolutionary origin. A phylogenetic study confirmed this possibility, positioning LipJ as a new member of the thermophilic family of bacterial lipases I.5. However, LipJ clusters in a clade close but separated from that of Geobacillus sp. thermophilic lipases. Comprehensive analysis of the cloned enzyme suggests a common origin of LipJ and other bacterial thermophilic lipases, and highlights the most probable divergent evolutionary pathway followed by LipJ, which during the harsh past times would have probably been a thermophilic enzyme, having lost these properties when the environment changed to more benign conditions.

  17. Contributions of a unique β-clamp to substrate recognition illuminates the molecular basis of exolysis in ferulic acid esterases.

    Science.gov (United States)

    Gruninger, Robert J; Cote, Chris; McAllister, Tim A; Abbott, D Wade

    2016-04-01

    Lignocellulosic biomass is a promising renewable resource; however, deconstruction of this material is still the rate-limiting step. Major obstacles in the biocatalytic turnover of lignocellulose are ester-linked decorations that prevent access to primary structural polysaccharides. Enzymes targeting these esters represent promising biotools for increasing bioconversion efficiency. Ruminant livestock are unique in their ability to degrade lignocellulose through the action of their gut microbiome. The anaerobic fungi (phylum Neocallimastigomycota) are key members of this ecosystem that express a large repertoire of carbohydrate-active enzymes (CAZymes) with little sequence identity with characterized CAZymes [Lombard, Golaconda, Drula, Coutinho and Henrissat (2014) Nucleic Acids Res. 42: , D490-D495]. We have identified a carbohydrate esterase family 1 (CE1) ferulic acid esterase (FAE) belonging to Anaeromyces mucronatus(AmCE1/Fae1a), and determined its X-ray structure in both the presence [1.55 Å (1 Å=0.1 nm)] and absence (1.60 Å) of ferulic acid. AmCE1 adopts an α/β-hydrolase fold that is structurally conserved with bacterial FAEs, and possesses a unique loop, termed the β-clamp, that encloses the ligand. Isothermal titration calorimetry reveals that substrate binding is driven by enthalpic contributions, which overcomes a large entropic penalty. A comparative analysis of AmCE1 with related enzymes has uncovered the apparent structural basis for differential FAE activities targeting cross-linking ferulic acid conjugates compared with terminal decorations. Based on comparisons to structurally characterized FAEs, we propose that the β-clamp may define the structural basis of exolytic activities in FAEs. This provides a structure-based tool for predicting exolysis and endolysis in CE1. These insights hold promise for rationally identifying enzymes tailored for bioconversion of biomass with variations in cell wall composition. © 2016 Authors; published by

  18. Plasma esterases in the tegu lizard Tupinambis merianae (Reptilia, Teiidae): impact of developmental stage, sex, and organophosphorus in vitro exposure.

    Science.gov (United States)

    Basso, Agustín; Attademo, Andrés M; Lajmanovich, Rafael C; Peltzer, Paola M; Junges, Celina; Cabagna, Mariana C; Fiorenza, Gabriela S; Sanchez-Hernandez, Juan Carlos

    2012-01-01

    In this study, we determined normal serum butyrylcholinesterase (BChE) and carboxylesterase (CbE) activities in Tupinambis merianae in order to obtain reference values for organophosphorus pesticide monitoring. Forty-two T. merianae individuals were grouped by sex and size to identify potential differences in their enzyme levels to allow for proper representation of normal values for females, males, juveniles, and hatchlings. Mean CbE was determined using two model substrates: alpha-naphtylacetate (α-NA) and p-nitrophenyl valerate (4-NPV). BChE and CbE sensitivity to malaoxon (Mx) was also evaluated as well as the possibility of BChE reactivation with pyridine-2-aldoxime methochloride (2-PAM). Mean adult females' BChE was significantly higher than adult males, juveniles, and hatchlings. No significant differences were found between groups regarding CbE. CbE (4-NPV) activity showed slightly negative correlation with lizard snout-vent length, while BChE and CbE (α-NA) showed no correlation with body size. Apparent IC(50) values for BChE and CbE (α-NA) suggested different sensitivities among groups. CbE (4-NPV) could not be inhibited. All Mx-inhibited groups treated with 2-PAM in a final concentration of 2.8 mM showed clear signs of reactivation. In conclusion, the results demonstrate that (1) plasma esterase activity did not vary with age and sex, except for BChE activity, and (2) because biological and environmental variables could be confounding factors in the response of plasma cholinesterases, complementary biomarkers like CbE inhibition and oxime-induced reactivation of esterases are strongly recommended.

  19. Rational design of a carboxylic esterase RhEst1 based on computational analysis of substrate binding.

    Science.gov (United States)

    Chen, Qi; Luan, Zheng-Jiao; Yu, Hui-Lei; Cheng, Xiaolin; Xu, Jian-He

    2015-11-01

    A new carboxylic esterase RhEst1 which catalyzes the hydrolysis of (S)-(+)-2,2-dimethylcyclopropanecarboxylate (S-DmCpCe), the key chiral building block of cilastatin, was identified and subsequently crystallized in our previous work. Mutant RhEst1A147I/V148F/G254A was found to show a 5-fold increase in the catalytic activity. In this work, molecular dynamic simulations were performed to elucidate the molecular determinant of the enzyme activity. Our simulations show that the substrate binds much more strongly in the A147I/V148F/G254A mutant than in wild type, with more hydrogen bonds formed between the substrate and the catalytic triad and the oxyanion hole. The OH group of the catalytic residue Ser101 in the mutant is better positioned to initiate the nucleophilic attack on S-DmCpCe. Interestingly, the "170-179" loop which is involved in shaping the catalytic sites and facilitating the product release shows remarkable dynamic differences in the two systems. Based on the simulation results, six residues were identified as potential "hot-spots" for further experimental testing. Consequently, the G126S and R133L mutants show higher catalytic efficiency as compared with the wild type. This work provides molecular-level insights into the substrate binding mechanism of carboxylic esterase RhEst1, facilitating future experimental efforts toward developing more efficient RhEst1 variants for industrial applications. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Biomonitoring of ecosystem degradation caused by CPO waste of Mentaya River in Central Kalimantan use of esterase isozyme electromorph method

    Directory of Open Access Journals (Sweden)

    PRABANG SETYONO

    2008-07-01

    Full Text Available The impact of CPO (Crude Palm Oil dock activity in Mentaya River of Central Borneo caused degradation of ecosystem, particularly on both mangrove and macrozoobenthos community. One of methods used for monitoring of ecosystem degradation was to determine species that were still survive under the polluted conditions. These survival species were assumed to synthesize alloenzyme that can be used as indicator. Alloenzyme was synthesized as an effort of adaptation processes toward environmental pressures caused by CPO spill on Mentaya River. Alloenzyme would be expressed as phenotypic and genotypic adaptation processes or phenotypic plasticity. Research was carried out, consisted of field research included collecting sample and environmental data (oil content, temperature, pH, electric conductivity and redox potential, and laboratory research included series analysis of water quality (DO, BOD, COD, pH, TSS, TDS and also alloenzyme content of Soneratia caseolaris L. and Macrobrachium rosenbergii de Man. The alloenzyme of root and leaves mangrove and prawn’s hepatopancreas was analyzed using Spencer starch gel electrophoresis modified method of exposed on sucrose solution. Separated components of alloenzyme were detected by special staining for Esterase isozyme. The results revealed that Soneratia caseolaris L. and Macrobrachium rosenbergii de Man were bioindicator organisms for the polluted site by oil spills from CPO loading activities. The polluted river water by oil spill from CPO activities decreased redox potential, DO, increased oil content, DHL, water temperature, pH sediment, pH water, TDS, BOD, COD, TSS. Gel electrophoretical analysis demonstrated that Mangrove Soneratia caseolaris synthesized alloenzyme consisted of complex enzymes such as EST in its root and leave cells. Those enzymes were nearly similar to those of Macrobrachium rosenbergii. The oil spill from CPO have ester bonding so its adaptation mechanism with release Esterase

  1. Ghosts of Milky Way's past: the globular cluster ESO 37-1 (E 3)

    Science.gov (United States)

    de la Fuente Marcos, R.; de la Fuente Marcos, C.; Moni Bidin, C.; Ortolani, S.; Carraro, G.

    2015-09-01

    Context. In the Milky Way, most globular clusters are highly conspicuous objects that were found centuries ago. However, a few dozen of them are faint, sparsely populated systems that were identified largely during the second half of the past century. One of the faintest is ESO 37-1 (E 3) and as such it remains poorly studied, with no spectroscopic observations published so far although it was discovered in 1976. Aims: We investigate the globular cluster E 3 in an attempt to better constrain its fundamental parameters. Spectroscopy of stars in the field of E 3 is shown here for the first time. Methods: Deep, precise VI CCD photometry of E 3 down to V ~ 26 mag is presented and analysed. Low-resolution, medium signal-to-noise ratio spectra of nine candidate members are studied to derive radial velocity and metallicity. Proper motions from the UCAC4 catalogue are used to explore the kinematics of the bright members of E 3. Results: Isochrone fitting indicates that E 3 is probably very old, with an age of about 13 Gyr; its distance from the Sun is nearly 10 kpc. It is also somewhat metal rich with [Fe/H] = -0.7. Regarding its kinematics, our tentative estimate for the proper motions is (μα cosδ,μδ) = (-7.0 ± 0.8, 3.5 ± 0.3) mas yr-1 (or a tangential velocity of 382 ± 79 km s-1) and for the radial velocity 45 ± 5 km s-1 in the solar rest frame. Conclusions: E 3 is one of the most intriguing globular clusters in the Galaxy. Having an old age and being metal rich is clearly a peculiar combination, only seen in a handful of objects like the far more conspicuous NGC 104 (47 Tucanae). In addition, its low luminosity and sparse population make it a unique template for the study of the final evolutionary phases in the life of a star cluster. Unfortunately, E 3 is among the most elusive and challenging known globular clusters because field contamination severely hampers spectroscopic studies. This research note is based on observations made with the ESO VLT at the

  2. Light Distribution in the E3 and E4 Scintillation Counters of the ATLAS Tile Calorimeter

    CERN Document Server

    Hsu, Catherine

    2013-01-01

    The Tile Calorimeter (TileCal) of the ATLAS experiment is an important component of the ATLAS calorimetry because they play a crucial role in the search for new particles. The E3 and E4 are crack scintillators of TileCal that extend into the gap region between the EM barrel and EM endcaps. They thus sample the energy of the EM showers produced by particles interacting with the dead material in the EM calorimeters and with the inner detector cables. This project focuses on the study of the light collection uniformity in the E3 and E4 scintillating tiles using low energy electrons as the ionising particles. It is important to have uniform light response in the tiles because it would ensure a good energy resolution for the dead region. However, many factors affect the uniform light collection within the scintillating tiles.

  3. Lunar Penetrating Radar onboard the Chang'e-3 mission

    International Nuclear Information System (INIS)

    Fang Guang-You; Zhou Bin; Ji Yi-Cai; Zhang Qun-Ying; Shen Shao-Xiang; Li Yu-Xi; Guan Hong-Fei; Tang Chuan-Jun; Gao Yun-Ze; Lu Wei; Ye Sheng-Bo; Han Hai-Dong; Zheng Jin; Wang Shu-Zhi

    2014-01-01

    Lunar Penetrating Radar (LPR) is one of the important scientific instruments onboard the Chang'e-3 spacecraft. Its scientific goals are the mapping of lunar regolith and detection of subsurface geologic structures. This paper describes the goals of the mission, as well as the basic principles, design, composition and achievements of the LPR. Finally, experiments on a glacier and the lunar surface are analyzed

  4. Proteolytic regulation of metabolic enzymes by E3 ubiquitin ligase complexes: lessons from yeast.

    Science.gov (United States)

    Nakatsukasa, Kunio; Okumura, Fumihiko; Kamura, Takumi

    2015-01-01

    Eukaryotic organisms use diverse mechanisms to control metabolic rates in response to changes in the internal and/or external environment. Fine metabolic control is a highly responsive, energy-saving process that is mediated by allosteric inhibition/activation and/or reversible modification of preexisting metabolic enzymes. In contrast, coarse metabolic control is a relatively long-term and expensive process that involves modulating the level of metabolic enzymes. Coarse metabolic control can be achieved through the degradation of metabolic enzymes by the ubiquitin-proteasome system (UPS), in which substrates are specifically ubiquitinated by an E3 ubiquitin ligase and targeted for proteasomal degradation. Here, we review select multi-protein E3 ligase complexes that directly regulate metabolic enzymes in Saccharomyces cerevisiae. The first part of the review focuses on the endoplasmic reticulum (ER) membrane-associated Hrd1 and Doa10 E3 ligase complexes. In addition to their primary roles in the ER-associated degradation pathway that eliminates misfolded proteins, recent quantitative proteomic analyses identified native substrates of Hrd1 and Doa10 in the sterol synthesis pathway. The second part focuses on the SCF (Skp1-Cul1-F-box protein) complex, an abundant prototypical multi-protein E3 ligase complex. While the best-known roles of the SCF complex are in the regulation of the cell cycle and transcription, accumulating evidence indicates that the SCF complex also modulates carbon metabolism pathways. The increasing number of metabolic enzymes whose stability is directly regulated by the UPS underscores the importance of the proteolytic regulation of metabolic processes for the acclimation of cells to environmental changes.

  5. A bacterial E3 ubiquitin ligase targets a host protein kinase to disrupt plant immunity.

    Science.gov (United States)

    Rosebrock, Tracy R; Zeng, Lirong; Brady, Jennifer J; Abramovitch, Robert B; Xiao, Fangming; Martin, Gregory B

    2007-07-19

    Many bacterial pathogens of plants and animals use a type III secretion system to deliver diverse virulence-associated 'effector' proteins into the host cell. The mechanisms by which these effectors act are mostly unknown; however, they often promote disease by suppressing host immunity. One type III effector, AvrPtoB, expressed by the plant pathogen Pseudomonas syringae pv. tomato, has a carboxy-terminal domain that is an E3 ubiquitin ligase. Deletion of this domain allows an amino-terminal region of AvrPtoB (AvrPtoB(1-387)) to be detected by certain tomato varieties leading to immunity-associated programmed cell death. Here we show that a host kinase, Fen, physically interacts with AvrPtoB(1-387 )and is responsible for activating the plant immune response. The AvrPtoB E3 ligase specifically ubiquitinates Fen and promotes its degradation in a proteasome-dependent manner. This degradation leads to disease susceptibility in Fen-expressing tomato lines. Various wild species of tomato were found to exhibit immunity in response to AvrPtoB(1-387 )and not to full-length AvrPtoB. Thus, by acquiring an E3 ligase domain, AvrPtoB has thwarted a highly conserved host resistance mechanism.

  6. Quantum phase transition in the U(4) vibron model and the E(3) symmetry

    International Nuclear Information System (INIS)

    Zhang Yu; Hou Zhanfeng; Chen Huan; Wei Haiqing; Liu Yuxin

    2008-01-01

    We study the details of the U(3)-O(4) quantum phase transition in the U(4) vibron model. Both asymptotic analysis in the classical limit and rigorous calculations for finite boson number systems indicate that a second-order phase transition is still there even for the systems with boson number N ranging from tens to hundreds. Two kinds of effective order parameters, including E1 transition ratios B(E1:2 1 →1 1 )/B(E1:1 1 →0 1 ) and B(E1:0 2 →1 1 )/B(E1:1 1 →0 1 ), and the energy ratios E 2 1 /E 0 2 and E 3 1 /E 0 2 are proposed to identify the second-order phase transition in experiments. We also found that the critical point of phase transition can be approximately described by the E(3) symmetry, which persists even for moderate N∼10 protected by the scaling behaviors of quantities at the critical point. In addition, a possible empirical example exhibiting roughly the E(3) symmetry is discussed

  7. HSV-1 ICP0: An E3 Ubiquitin Ligase That Counteracts Host Intrinsic and Innate Immunity

    Directory of Open Access Journals (Sweden)

    Mirna Perusina Lanfranca

    2014-05-01

    Full Text Available The herpes simplex virus type 1 (HSV-1 encoded E3 ubiquitin ligase, infected cell protein 0 (ICP0, is required for efficient lytic viral replication and regulates the switch between the lytic and latent states of HSV-1. As an E3 ubiquitin ligase, ICP0 directs the proteasomal degradation of several cellular targets, allowing the virus to counteract different cellular intrinsic and innate immune responses. In this review, we will focus on how ICP0’s E3 ubiquitin ligase activity inactivates the host intrinsic defenses, such as nuclear domain 10 (ND10, SUMO, and the DNA damage response to HSV-1 infection. In addition, we will examine ICP0’s capacity to impair the activation of interferon (innate regulatory mediators that include IFI16 (IFN γ-inducible protein 16, MyD88 (myeloid differentiation factor 88, and Mal (MyD88 adaptor-like protein. We will also consider how ICP0 allows HSV-1 to evade activation of the NF-κB (nuclear factor kappa B inflammatory signaling pathway. Finally, ICP0’s paradoxical relationship with USP7 (ubiquitin specific protease 7 and its roles in intrinsic and innate immune responses to HSV-1 infection will be discussed.

  8. Construction of new operation interface for the LABIHS simulator using the ELIPSE E3 studio software

    Energy Technology Data Exchange (ETDEWEB)

    Augusto, Silas C.; Oliveira, Mauro V., E-mail: silas@ien.gov.b, E-mail: mvitor@ien.gov.b [Instituto de Engenharia Nuclear (IEN/CNEN-RJ), Rio de Janeiro, RJ (Brazil)

    2011-07-01

    The Human-System Interface Laboratory (LABIHS), located at the Instituto de Engenharia Nuclear (IEN), has a compact simulator that simulate the processes of a pressurized water reactor nuclear power plant of 930 MWe of power. This simulator is composed by a HP-UX workstation computer, where the simulation software runs, and a set of computer stations, that represent an advanced control room, where the simulator is operated by software control panels that represent several systems of the simulated nuclear power plant. The current HSIs for the LABIHS simulator was built using iLog software tool. The development of new human-system interfaces (HSIs) for the simulator is one of the research fields of LABIHS. This paper presents the screen components development process for a new HSI for the LABIHS simulator, using the software Elipse{sup TM} E3 Studio. These new components developed using the E3 Studio are similar to the ones used in the current simulator interface. The article shows some comparisons between the component and screen development with Elipse{sup TM} E3 Studio processes and using iLog Studio. (author)

  9. Construction of new operation interface for the LABIHS simulator using the ELIPSE E3 studio software

    International Nuclear Information System (INIS)

    Augusto, Silas C.; Oliveira, Mauro V.

    2011-01-01

    The Human-System Interface Laboratory (LABIHS), located at the Instituto de Engenharia Nuclear (IEN), has a compact simulator that simulate the processes of a pressurized water reactor nuclear power plant of 930 MWe of power. This simulator is composed by a HP-UX workstation computer, where the simulation software runs, and a set of computer stations, that represent an advanced control room, where the simulator is operated by software control panels that represent several systems of the simulated nuclear power plant. The current HSIs for the LABIHS simulator was built using iLog software tool. The development of new human-system interfaces (HSIs) for the simulator is one of the research fields of LABIHS. This paper presents the screen components development process for a new HSI for the LABIHS simulator, using the software Elipse TM E3 Studio. These new components developed using the E3 Studio are similar to the ones used in the current simulator interface. The article shows some comparisons between the component and screen development with Elipse TM E3 Studio processes and using iLog Studio. (author)

  10. Identification of Arabidopsis MYB56 as a novel substrate for CRL3(BPM) E3 ligases.

    Science.gov (United States)

    Chen, Liyuan; Bernhardt, Anne; Lee, JooHyun; Hellmann, Hanjo

    2015-02-01

    Controlled stability of proteins is a highly efficient mechanism to direct diverse processes in living cells. A key regulatory system for protein stability is given by the ubiquitin proteasome pathway, which uses E3 ligases to mark specific proteins for degradation. In this work, MYB56 is identified as a novel target of a CULLIN3 (CUL3)-based E3 ligase. Its stability depends on the presence of MATH-BTB/POZ (BPM) proteins, which function as substrate adaptors to the E3 ligase. Genetic studies have indicated that MYB56 is a negative regulator of flowering, while BPMs positively affect this developmental program. The interaction between BPMs and MYB56 occurs at the promoter of FLOWERING LOCUS T (FT), a key regulator in initiating flowering in Arabidopsis, and results in instability of MYB56. Overall the work establishes MYB transcription factors as substrates of BPM proteins, and provides novel information on components that participate in controlling flowering time in plants. Copyright © 2015 The Author. Published by Elsevier Inc. All rights reserved.

  11. Identification of Arabidopsis MYB56 as a novel substrate for CRL3BPM E3 ligases.

    Science.gov (United States)

    Chen, Liyuan; Bernhardt, Anne; Lee, JooHyun; Hellmann, Hanjo

    2014-10-24

    Controlled stability of proteins is a highly efficient mechanism to direct diverse processes in living cells. A key regulatory system for protein stability is given by the ubiquitin proteasome pathway, which uses E3 ligases to mark specific proteins for degradation. In this work MYB56 is identified as a novel target of a CULLIN3 (CUL3)-based E3 ligase. Its stability depends on the presence of MATH-BTB/POZ (BPM) proteins, which function as substrate adaptors to the E3 ligase. Genetic studies pointed out that MYB56 is a negative regulator of flowering, while BPMs positively affect this developmental program. The interaction between BPMs and MYB56 occurs at the promoter of FLOWERING LOCUS T (FT), a key regulator in initiating flowering in Arabidopsis, and results in instability of MYB56. Overall the work establishes MYB transcription factors as substrates of BPM proteins, and provides novel information on components that participate in controlling the flowering time point in plants. © The Author 2014. Published by the Molecular Plant Shanghai Editorial Office in association with Oxford University Press on behalf of CSPB and IPPE, SIBS, CAS.

  12. Functional analysis of NopM, a novel E3 ubiquitin ligase (NEL domain effector of Rhizobium sp. strain NGR234.

    Directory of Open Access Journals (Sweden)

    Da-Wei Xin

    Full Text Available Type 3 effector proteins secreted via the bacterial type 3 secretion system (T3SS are not only virulence factors of pathogenic bacteria, but also influence symbiotic interactions between nitrogen-fixing nodule bacteria (rhizobia and leguminous host plants. In this study, we characterized NopM (nodulation outer protein M of Rhizobium sp. strain NGR234, which shows sequence similarities with novel E3 ubiquitin ligase (NEL domain effectors from the human pathogens Shigella flexneri and Salomonella enterica. NopM expressed in Escherichia coli, but not the non-functional mutant protein NopM-C338A, showed E3 ubiquitin ligase activity in vitro. In vivo, NopM, but not inactive NopM-C338A, promoted nodulation of the host plant Lablab purpureus by NGR234. When NopM was expressed in yeast, it inhibited mating pheromone signaling, a mitogen-activated protein (MAP kinase pathway. When expressed in the plant Nicotiana benthamiana, NopM inhibited one part of the plant's defense response, as shown by a reduced production of reactive oxygen species (ROS in response to the flagellin peptide flg22, whereas it stimulated another part, namely the induction of defense genes. In summary, our data indicate the potential for NopM as a functional NEL domain E3 ubiquitin ligase. Our findings that NopM dampened the flg22-induced ROS burst in N. benthamiana but promoted defense gene induction are consistent with the concept that pattern-triggered immunity is split in two separate signaling branches, one leading to ROS production and the other to defense gene induction.

  13. GEM-E3: A computable general equilibrium model applied for Switzerland

    Energy Technology Data Exchange (ETDEWEB)

    Bahn, O. [Paul Scherrer Inst., CH-5232 Villigen PSI (Switzerland); Frei, C. [Ecole Polytechnique Federale de Lausanne (EPFL) and Paul Scherrer Inst. (Switzerland)

    2000-01-01

    The objectives of the European Research Project GEM-E3-ELITE, funded by the European Commission and coordinated by the Centre for European Economic Research (Germany), were to further develop the general equilibrium model GEM-E3 (Capros et al., 1995, 1997) and to conduct policy analysis through case studies. GEM-E3 is an applied general equilibrium model that analyses the macro-economy and its interaction with the energy system and the environment through the balancing of energy supply and demand, atmospheric emissions and pollution control, together with the fulfillment of overall equilibrium conditions. PSI's research objectives within GEM-E3-ELITE were to implement and apply GEM-E3 for Switzerland. The first objective required in particular the development of a Swiss database for each of GEM-E3 modules (economic module and environmental module). For the second objective, strategies to reduce CO{sub 2} emissions were evaluated for Switzerland. In order to develop the economic, PSI collaborated with the Laboratory of Applied Economics (LEA) of the University of Geneva and the Laboratory of Energy Systems (LASEN) of the Federal Institute of Technology in Lausanne (EPFL). The Swiss Federal Statistical Office (SFSO) and the Institute for Business Cycle Research (KOF) of the Swiss Federal Institute of Technology (ETH Zurich) contributed also data. The Swiss environmental database consists mainly of an Energy Balance Table and of an Emission Coefficients Table. Both were designed using national and international official statistics. The Emission Coefficients Table is furthermore based on know-how of the PSI GaBE Project. Using GEM-E3 Switzerland, two strategies to reduce the Swiss CO{sub 2} emissions were evaluated: a carbon tax ('tax only' strategy), and the combination of a carbon tax with the buying of CO{sub 2} emission permits ('permits and tax' strategy). In the first strategy, Switzerland would impose the necessary carbon tax to achieve

  14. GEM-E3: A computable general equilibrium model applied for Switzerland

    International Nuclear Information System (INIS)

    Bahn, O.; Frei, C.

    2000-01-01

    The objectives of the European Research Project GEM-E3-ELITE, funded by the European Commission and coordinated by the Centre for European Economic Research (Germany), were to further develop the general equilibrium model GEM-E3 (Capros et al., 1995, 1997) and to conduct policy analysis through case studies. GEM-E3 is an applied general equilibrium model that analyses the macro-economy and its interaction with the energy system and the environment through the balancing of energy supply and demand, atmospheric emissions and pollution control, together with the fulfillment of overall equilibrium conditions. PSI's research objectives within GEM-E3-ELITE were to implement and apply GEM-E3 for Switzerland. The first objective required in particular the development of a Swiss database for each of GEM-E3 modules (economic module and environmental module). For the second objective, strategies to reduce CO 2 emissions were evaluated for Switzerland. In order to develop the economic, PSI collaborated with the Laboratory of Applied Economics (LEA) of the University of Geneva and the Laboratory of Energy Systems (LASEN) of the Federal Institute of Technology in Lausanne (EPFL). The Swiss Federal Statistical Office (SFSO) and the Institute for Business Cycle Research (KOF) of the Swiss Federal Institute of Technology (ETH Zurich) contributed also data. The Swiss environmental database consists mainly of an Energy Balance Table and of an Emission Coefficients Table. Both were designed using national and international official statistics. The Emission Coefficients Table is furthermore based on know-how of the PSI GaBE Project. Using GEM-E3 Switzerland, two strategies to reduce the Swiss CO 2 emissions were evaluated: a carbon tax ('tax only' strategy), and the combination of a carbon tax with the buying of CO 2 emission permits ('permits and tax' strategy). In the first strategy, Switzerland would impose the necessary carbon tax to achieve the reduction target, and use the tax

  15. Purification and general properties of pectin methyl esterase from Curvularia inaequalis NRRL 13884 in solid state culture using orange peels as an inducer.

    Science.gov (United States)

    Afifi, A F; Fawzi, E M; Foaad, M A

    2002-01-01

    Pectin methyl esterase (PME) [E.C.3. 1.1.11] production by Curvularia inaequalis (Shear) Boedijn NRRL 13884 was investigated using solid-state culture. The highest level of extracellular pectin methyl esterase was detected with orange peels as an inducing substrate and as a sole carbon source. The enzyme was partially purified using Sephadex G-100 and DEAE-Cellulose column chromatography. It was purified about 40 fold with optimum activity at pH 4.4 and 45 degrees C. The enzyme was activated by Co++, Mg++, Na+, whereas it was slightly activated in the presence of Cu++, K+, Mn++, Zn++. On the other hand Ag++, Ca++ and Hg++ inhibited the activity of the enzyme. The Km was calculated to be 0.52 mM.

  16. Notes on electropherograms of eye-lens, muscle proteins and zymograms of muscle esterases of fish collected during the first Brazilian expedition to the Antarctica

    Directory of Open Access Journals (Sweden)

    Van Ngan Phan

    1985-01-01

    Full Text Available A preliminary study was carried out on electropherograms of eye-lens, muscle proteins and zymograms of muscle esterases of ten Notothenia larseni, six Notothenia nudifrons and one lanternfish, Electrona antarctica. The fish were collected by the R/V "Prof. W. Besnard" of the Institute of Oceanography, University of São Paulo, during the First Brazilian Expedition to Antarctica. Eye-lens proteins were analysed on cellulose acetate membrane, muscle proteins and esterases on gel of polyaorylamide. Eye-lens proteins showed three types of electropherograms for N. larseni, and two types for N. nudifrons. One of the electropherograms of N. larseni can be readily distinguished from those of N. nudifrons. Electropherograms of muscle proteins of N. larseni and N. nudifrons are very similar and, consist of sixteen to seventeen fractions. Electropherograms of muscle proteins of N. larseni are severely affected by the conservation of the extracts overnight under -20ºC. All N. nudifrons were of the same zymograms of esterases while those of N. larseni varied. Electropherograms of eye-lens and muscle proteins as well as zymograms of esterases of the lanternfish are different from those of nototheniids.Foi realizado um estudo preliminar sobre eletroferogramas de proteínas de cristalino e de músculo esquelético, e zimogramas de esterases de músculo esquelético de dez Notothenia larseni, seis Notothenia nudifrons e de um peixe-lanterna, Electrona antarctica. Os peixes foram coletados pelo N/Oc. "Prof. W. Besnard" do Instituto Oceanográfico da Universidade de São Paulo durante a I Expedição Brasileira à Antártica. As proteinas do cristalino foram analisadas em membranas de acetato de celulose, enquanto que as proteínas e esterases do músculo esquelético, em gel de poliacrilamida. As proteínas do cristalino apresentam três tipos distintos de eletroferogramas para N. larseni, e dois para N. nudifrons. Um dos eletroferogramas de N. larseni, pode ser

  17. Preliminary X-ray analysis of twinned crystals of the Q88Y25-Lacpl esterase from Lactobacillus plantarum WCFS1

    International Nuclear Information System (INIS)

    Álvarez, Yanaisis; Esteban-Torres, María; Acebrón, Iván; Rivas, Blanca de las; Muñoz, Rosario; Martínez-Ripoll, Martín; Mancheño, José M.

    2011-01-01

    The Q88Y25-Lacpl esterase from L. plantarum WCFS1 has been recombinantly expressed, purified and crystallized. A native diffraction data set has been collected to 2.24 Å resolution. Q88Y25-Lacpl is an esterase produced by the lactic acid bacterium Lactobacillus plantarum WCFS1 that shows amino-acid sequence similarity to carboxylesterases from the hormone-sensitive lipase family, in particular the AFEST esterase from the archaeon Archaeoglobus fulgidus and the hyperthermophilic esterase EstEI isolated from a metagenomic library. N-terminally His 6 -tagged Q88Y25-Lacpl has been overexpressed in Escherichia coli BL21 (DE3) cells, purified and crystallized at 291 K using the hanging-drop vapour-diffusion method. Mass spectrometry was used to determine the purity and homogeneity of the enzyme. Crystals of His 6 -tagged Q88Y25-Lacpl were prepared in a solution containing 2.8 M sodium acetate trihydrate pH 7.0. X-ray diffraction data were collected to 2.24 Å resolution on beamline ID29 at the ESRF. The apparent crystal point group was 422; however, initial global analysis of the intensity statistics (data processed with high symmetry in space group I422) and subsequent tests on data processed with low symmetry (space group I4) showed that the crystals were almost perfectly merohedrally twinned. Most probably, the true space group is I4, with unit-cell parameters a = 169.05, b = 169.05, c = 183.62 Å

  18. The Arabidopsis E3 Ubiquitin Ligase HOS1 Negatively Regulates CONSTANS Abundance in the Photoperiodic Control of Flowering[W

    Science.gov (United States)

    Lazaro, Ana; Valverde, Federico; Piñeiro, Manuel; Jarillo, Jose A.

    2012-01-01

    The Arabidopsis thaliana early in short days6 (esd6) mutant was isolated in a screen for mutations that accelerate flowering time. Among other developmental alterations, esd6 displays early flowering in both long- and short-day conditions. Fine mapping of the mutation showed that the esd6 phenotype is caused by a lesion in the HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES1 (HOS1) locus, which encodes a RING finger–containing E3 ubiquitin ligase. The esd6/hos1 mutation causes decreased FLOWERING LOCUS C expression and requires CONSTANS (CO) protein for its early flowering phenotype under long days. Moreover, CO and HOS1 physically interact in vitro and in planta, and HOS1 regulates CO abundance, particularly during the daylight period. Accordingly, hos1 causes a shift in the regular long-day pattern of expression of FLOWERING LOCUS T (FT) transcript, starting to rise 4 h after dawn in the mutant. In addition, HOS1 interacts synergistically with CONSTITUTIVE PHOTOMORPHOGENIC1, another regulator of CO protein stability, in the regulation of flowering time. Taken together, these results indicate that HOS1 is involved in the control of CO abundance, ensuring that CO activation of FT occurs only when the light period reaches a certain length and preventing precocious flowering in Arabidopsis. PMID:22408073

  19. The E3 Ubiquitin Ligase MIB-1 Is Necessary To Form the Nuclear Halo in Caenorhabditis elegans Sperm.

    Science.gov (United States)

    Herrera, Leslie A; Starr, Daniel A

    2018-05-18

    Unlike the classical nuclear envelope with two membranes found in other eukaryotic cells, most nematode sperm nuclei are not encapsulated by membranes. Instead, they are surrounded by a nuclear halo of unknown composition. How the halo is formed and regulated is unknown. We used forward genetics to identify molecular lesions behind three classical fer (fertilization defective) mutations that disrupt the ultrastructure of the Caenorhabditis elegans sperm nuclear halo. We found fer-2 and fer-4 alleles to be nonsense mutations in mib-1. fer-3 was caused by a nonsense mutation in eri-3 GFP::MIB-1 was expressed in the germline during early spermatogenesis, but not in mature sperm. mib-1 encodes a conserved E3 ubiquitin ligase homologous to vertebrate Mib1 and Mib2, which function in Notch signaling. Here, we show that mib-1 is important for male sterility and is involved in the regulation or formation of the nuclear halo during nematode spermatogenesis. Copyright © 2018, G3: Genes, Genomes, Genetics.

  20. Structure of the Human FANCL RING-Ube2T Complex Reveals Determinants of Cognate E3-E2 Selection

    Science.gov (United States)

    Hodson, Charlotte; Purkiss, Andrew; Miles, Jennifer Anne; Walden, Helen

    2014-01-01

    Summary The combination of an E2 ubiquitin-conjugating enzyme with an E3 ubiquitin-ligase is essential for ubiquitin modification of a substrate. Moreover, the pairing dictates both the substrate choice and the modification type. The molecular details of generic E3-E2 interactions are well established. Nevertheless, the determinants of selective, specific E3-E2 recognition are not understood. There are ∼40 E2s and ∼600 E3s giving rise to a possible ∼24,000 E3-E2 pairs. Using the Fanconi Anemia pathway exclusive E3-E2 pair, FANCL-Ube2T, we report the atomic structure of the FANCL RING-Ube2T complex, revealing a specific and extensive network of additional electrostatic and hydrophobic interactions. Furthermore, we show that these specific interactions are required for selection of Ube2T over other E2s by FANCL. PMID:24389026

  1. Expression of mung bean pectin acetyl esterase in potato tubers: effect on acetylation of cell wall polymers and tuber mechanical properties.

    Science.gov (United States)

    Orfila, Caroline; Dal Degan, Florence; Jørgensen, Bodil; Scheller, Henrik Vibe; Ray, Peter M; Ulvskov, Peter

    2012-07-01

    A mung bean (Vigna radiata) pectin acetyl esterase (CAA67728) was heterologously expressed in tubers of potato (Solanum tuberosum) under the control of the granule-bound starch synthase promoter or the patatin promoter in order to probe the significance of O-acetylation on cell wall and tissue properties. The recombinant tubers showed no apparent macroscopic phenotype. The enzyme was recovered from transgenic tubers using a high ionic strength buffer and the extract was active against a range of pectic substrates. Partial in vivo de-acetylation of cell wall polysaccharides occurred in the transformants, as shown by a 39% decrease in the degree of acetylation (DA) of tuber cell wall material (CWM). Treatment of CWM using a combination of endo-polygalacturonase and pectin methyl esterase extracted more pectin polymers from the transformed tissue compared to wild type. The largest effect of the pectin acetyl esterase (68% decrease in DA) was seen in the residue from this extraction, suggesting that the enzyme is preferentially active on acetylated pectin that is tightly bound to the cell wall. The effects of acetylation on tuber mechanical properties were investigated by tests of failure under compression and by determination of viscoelastic relaxation spectra. These tests suggested that de-acetylation resulted in a stiffer tuber tissue and a stronger cell wall matrix, as a result of changes to a rapidly relaxing viscoelastic component. These results are discussed in relation to the role of pectin acetylation in primary cell walls and its implications for industrial uses of potato fibres.

  2. Refractory Abdominal Pain in a Patient with Chronic Lymphocytic Leukemia: Be Wary of Acquired Angioedema due to C1 Esterase Inhibitor Deficiency

    Directory of Open Access Journals (Sweden)

    Abdullateef Abdulkareem

    2018-01-01

    Full Text Available Acquired angioedema due to C1 inhibitor deficiency (C1INH-AAE is a rare and potentially fatal syndrome of bradykinin-mediated angioedema characterized by episodes of angioedema without urticaria. It typically manifests with nonpitting edema of the skin and edema in the gastrointestinal (GI tract mucosa or upper airway. Edema of the upper airway and tongue may lead to life-threatening asphyxiation. C1INH-AAE is typically under-diagnosed because of its rarity and its propensity to mimic more common abdominal conditions and allergic reactions. In this article, we present the case of a 62-year-old male with a history of recently diagnosed chronic lymphocytic leukemia (CLL who presented to our hospital with recurrent abdominal pain, initially suspected to have Clostridium difficile colitis and diverticulitis. He received a final diagnosis of acquired angioedema due to C1 esterase inhibitor deficiency due to concomitant symptoms of lip swelling, cutaneous nonpitting edema of his lower extremities, and complement level deficiencies. He received acute treatment with C1 esterase replacement and icatibant and was maintained on C1 esterase infusions. He also underwent chemotherapy for his underlying CLL and did not experience further recurrence of his angioedema.

  3. Novel Feruloyl Esterase from Lactobacillus fermentum NRRL B-1932 and Analysis of the Recombinant Enzyme Produced in Escherichia coli.

    Science.gov (United States)

    Liu, Siqing; Bischoff, Kenneth M; Anderson, Amber M; Rich, Joseph O

    2016-09-01

    A total of 33 Lactobacillus strains were screened for feruloyl esterase (FE) activity using agar plates containing ethyl ferulate as the sole carbon source, and Lactobacillus fermentum NRRL B-1932 demonstrated the strongest FE activity among a dozen species showing a clearing zone on the opaque plate containing ethyl ferulate. FE activities were monitored using high-performance liquid chromatography with an acetonitrile-trifluoroacetic acid gradient. To produce sufficient purified FE from L. fermentum strain NRRL B-1932 (LfFE), the cDNA encoding LfFE (Lffae) was amplified and cloned by using available closely related genome sequences and overexpressed in Escherichia coli A 29.6-kDa LfFE protein was detected from the protein extract of E. coli BL21(pLysS) carrying pET28bLffae upon IPTG (isopropyl-β-d-thiogalactopyranoside) induction. The recombinant LfFE containing a polyhistidine tag was purified by nickel-nitrilotriacetic acid affinity resin. The purified LfFE showed strong activities against several artificial substrates, including p-nitrophenyl acetate and 4-methylumbelliferyl p-trimethylammoniocinnamate chloride. The optimum pH and temperature of the recombinant LfFE were around 6.5 and 37°C, respectively, as determined using either crude or purified recombinant LfFE. This study will be essential for the production of the LfFE in E. coli on a larger scale that could not be readily achieved by L. fermentum fermentation. The production of feruloyl esterase (FE) from Lactobacillus fermentum NRRL B-1932 reported in this study will have immense potential commercial applications not only in biofuel production but also in pharmaceutical, polymer, oleo chemical, cosmetic additive, and detergent industries, as well as human health-related applications, including food flavoring, functional foods, probiotic agents, preventive medicine, and animal feed. Given the essential role FE plays in the production of hydroxycinnamic acids and ferulic acid, plus the generally

  4. The Processing and Analysis of Lunar Penetrating Radar Channel-1 Data from Chang'E-3

    Directory of Open Access Journals (Sweden)

    Gao Yun-ze

    2015-10-01

    Full Text Available Lunar Penetrating Radar (LPR, which is one of the most important science payloads onboard the Chang'E-3 (CE-3 rover, is used to obtain electromagnetic image less than 100 m beneath the lunar surface. This paper describes the system composition and working mechanism of the LPR and presents a detailed analysis of its data. We investigated special signal-processing methods and present the result of channel-1 data. The result shows that the effective echo occurs at depths greater than 100 m. Moreover, an unusual reflection exists at depth of 40 m, which may be the boundary of two geological units beneath the lunar surface.

  5. Subsurface structures in the northern Mare Imbrium measured by Chang'E-3 and SELENE

    Science.gov (United States)

    Kumamoto, A.; Ishiyama, K.; Feng, J.

    2016-12-01

    Subsurface structures in the northern Mare Imbrium measured by Chang'E-3 and SELENE have been compared. In Chang'E-3 mission, subsurface radar sounding at (19.51W, 44.12N) was performed by Lunar Penetrating Radar (LPR) onboard the Yutu rover. The LPR was pulse radar operated at two frequencies: 60 MHz and 500 MHz. During its operation period from December 2013 to January 2014, the LPR observed subsurface echoes along the rover's track with total distance of 114 m. From the observation in 60 MHz, the subsurface echoes from buried regolith layers at depths of 35, 50, 140, 240, and 360 m were reported (Xiao et al., 2015). In SELENE mission, global subsurface radar sounding of the moon was performed by Lunar Radar Sounder (LRS) onboard the SELENE (Kaguya) spacecraft from the polar orbit with an altitude of 100 km. The LRS was chirp radar operated in a frequency range from 4-6 MHz. So the range resolution of LRS was 75 m in vacuum. During operation period from December 2007 to September 2008, subsurface echoes from all areas of the Moon was observed with a lateral resolution of 76 m. From the global observation, the subsurface echoes from the buried regolith layers in the neraside maria including Mare Imbrium at depths of several hundred meters were reported (Ono et al., 2009).In the present study, we focus on SELENE/LRS data obtained at (19.50W, 44.12N) which is the nearest to the Chang'E-3 landing site. While clear and large-scale subsurface reflectors, as found in Ono et al. (2009), are not found in it, we can identify some echo components from the depths of 140 ( 2000 ns), 240 ( 4000 ns), and 360 m ( 6000 ns). Further analyses utilizing high-resolution data from Chang'E-3/LPR and large-scale data from SELENE/LRS, we will be able to determine the thickness and large-scale structures of the buried regolith layers found by the both radars, and discuss their formation processes in volcanic history of Mare Imbrium.

  6. The Processing and Analysis of Lunar Penetrating Radar Channel-1 Data from Chang'E-3

    OpenAIRE

    Gao Yun-ze; Dong Ze-hua; Fang Guang-you; Ji Yi-cai; Zhou Bin

    2015-01-01

    Lunar Penetrating Radar (LPR), which is one of the most important science payloads onboard the Chang'E-3 (CE-3) rover, is used to obtain electromagnetic image less than 100 m beneath the lunar surface. This paper describes the system composition and working mechanism of the LPR and presents a detailed analysis of its data. We investigated special signal-processing methods and present the result of channel-1 data. The result shows that the effective echo occurs at depths greater than 100 m. Mo...

  7. High-spin yrast isomers in 211Rn and 212Rn with enhanced E3 decays

    International Nuclear Information System (INIS)

    Dracoulis, G.D.; Byrne, A.P.; Fabricius, B.

    1990-01-01

    New isomeric states with J π =69/2 + ,τ m = 13 (1) ns in 211 Rn and J π =33 - ,τ m = 7(1) ns in 212 Rn have been identified. They decay by enchanced E3 transitions with strengths of 33(3) and 43(6) single particle units to the known 63/2 - and 30 + isomers 211 Rn and 212 Rn, respectively. The excitation energies and transition strengths agree well with predictions of the multi-particle, octupole-vibration coupled model. 13 refs., 2 tabs., 3 figs

  8. IFT20 modulates ciliary PDGFRα signaling by regulating the stability of Cbl E3 ubiquitin ligases

    DEFF Research Database (Denmark)

    Schmid, Fabian Marc; Schou, Kenneth Bødtker; Vilhelm, Martin Juel

    2018-01-01

    ciliogenesis, and ciliary localization of the receptor is required for its appropriate ligand-mediated activation by PDGF-AA. However, the mechanisms regulating sorting of PDGFRα and feedback inhibition of PDGFRα signaling at the cilium are unknown. Here, we provide evidence that intraflagellar transport...... protein 20 (IFT20) interacts with E3 ubiquitin ligases c-Cbl and Cbl-b and is required for Cbl-mediated ubiquitination and internalization of PDGFRα for feedback inhibition of receptor signaling. In wild-type cells treated with PDGF-AA, c-Cbl becomes enriched in the cilium, and the receptor...

  9. Coplanar (e, 3e) differential cross-section of He atom

    CERN Document Server

    Choubisa, R; Sud, K K

    2003-01-01

    We present in this paper the results of our calculation of five-fold differential cross-section (FDCS) for (e, 3e) process on He atom in low momentum transfer and high electron impact energy in shake-off mechanism. The formalism has been developed in Born approximation using plane waves. Byron and Joachain as well as Le Sech and correlated BBK-type wave functions respectively for incident and scattered. bound and ejected electrons. The angular distribution of FDCS of our calculation is presented in various modes of coplanar geometry and comparison is made with the available experimental data. We observe that the present calculation is able to reproduce the trend of the experimental data. However, it differs in magnitude from the experiment. The present theory does not predict four-peak structure in summed mutual angle mode for lower excess ejected electron energies. We also discuss the importance of momentum transfer, post-collision interaction (PCI) and ion participation in the (e, 3e) process in constant th...

  10. No Evidence for Multiple Stellar Populations in the Low-mass Galactic Globular Cluster E 3

    Science.gov (United States)

    Salinas, Ricardo; Strader, Jay

    2015-08-01

    Multiple stellar populations are a widespread phenomenon among Galactic globular clusters. Even though the origin of the enriched material from which new generations of stars are produced remains unclear, it is likely that self-enrichment will be feasible only in clusters massive enough to retain this enriched material. We searched for multiple populations in the low mass (M˜ 1.4× {10}4 {M}⊙ ) globular cluster E3, analyzing SOAR/Goodman multi-object spectroscopy centered on the blue cyanogen (CN) absorption features of 23 red giant branch stars. We find that the CN abundance does not present the typical bimodal behavior seen in clusters hosting multistellar populations, but rather a unimodal distribution that indicates the presence of a genuine single stellar population, or a level of enrichment much lower than in clusters that show evidence for two populations from high-resolution spectroscopy. E3 would be the first bona fide Galactic old globular cluster where no sign of self-enrichment is found. Based on observations obtained at the Southern Astrophysical Research (SOAR) Telescope, which is a joint project of the Ministério da Ciência, Tecnologia, e Inovação (MCTI) da República Federativa do Brasil, the US National Optical Astronomy Observatory (NOAO), the University of North Carolina at Chapel Hill (UNC), and Michigan State University (MSU).

  11. Aurora Kinase A Promotes AR Degradation via the E3 Ligase CHIP.

    Science.gov (United States)

    Sarkar, Sukumar; Brautigan, David L; Larner, James M

    2017-08-01

    Reducing the levels of the androgen receptor (AR) is one of the most viable approaches to combat castration-resistant prostate cancer. Previously, we observed that proteasomal-dependent degradation of AR in response to 2-methoxyestradiol (2-ME) depends primarily on the E3 ligase C-terminus of HSP70-interacting protein (STUB1/CHIP). Here, 2-ME stimulation activates CHIP by phosphorylation via Aurora kinase A (AURKA). Aurora A kinase inhibitors and RNAi knockdown of Aurora A transcript selectively blocked CHIP phosphorylation and AR degradation. Aurora A kinase is activated by 2-ME in the S-phase as well as during mitosis, and phosphorylates CHIP at S273. Prostate cancer cells expressing an S273A mutant of CHIP have attenuated AR degradation upon 2-ME treatment compared with cells expressing wild-type CHIP, supporting the idea that CHIP phosphorylation by Aurora A activates its E3 ligase activity for the AR. These results reveal a novel 2-ME→Aurora A→CHIP→AR pathway that promotes AR degradation via the proteasome that may offer novel therapeutic opportunities for prostate cancer. Mol Cancer Res; 15(8); 1063-72. ©2017 AACR . ©2017 American Association for Cancer Research.

  12. Blocking an N-terminal acetylation–dependent protein interaction inhibits an E3 ligase

    Energy Technology Data Exchange (ETDEWEB)

    Scott, Daniel C.; Hammill, Jared T.; Min, Jaeki; Rhee, David Y.; Connelly, Michele; Sviderskiy, Vladislav O.; Bhasin, Deepak; Chen, Yizhe; Ong, Su-Sien; Chai, Sergio C.; Goktug, Asli N.; Huang, Guochang; Monda, Julie K.; Low, Jonathan; Kim, Ho Shin; Paulo, Joao A.; Cannon, Joe R.; Shelat, Anang A.; Chen, Taosheng; Kelsall, Ian R.; Alpi, Arno F.; Pagala, Vishwajeeth; Wang, Xusheng; Peng, Junmin; Singh , Bhuvanesh; Harper, J. Wade; Schulman, Brenda A.; Guy, R. Kip (MSKCC); (Dundee); (SJCH); (Harvard-Med); (MXPL)

    2017-06-05

    N-terminal acetylation is an abundant modification influencing protein functions. Because ~80% of mammalian cytosolic proteins are N-terminally acetylated, this modification is potentially an untapped target for chemical control of their functions. Structural studies have revealed that, like lysine acetylation, N-terminal acetylation converts a positively charged amine into a hydrophobic handle that mediates protein interactions; hence, this modification may be a druggable target. We report the development of chemical probes targeting the N-terminal acetylation–dependent interaction between an E2 conjugating enzyme (UBE2M or UBC12) and DCN1 (DCUN1D1), a subunit of a multiprotein E3 ligase for the ubiquitin-like protein NEDD8. The inhibitors are highly selective with respect to other protein acetyl-amide–binding sites, inhibit NEDD8 ligation in vitro and in cells, and suppress anchorage-independent growth of a cell line with DCN1 amplification. Overall, our data demonstrate that N-terminal acetyl-dependent protein interactions are druggable targets and provide insights into targeting multiprotein E2–E3 ligases.

  13. Diagramming Transactive Building Business Cases: Using Principles of e3 Value to Document Valuation Studies

    Energy Technology Data Exchange (ETDEWEB)

    Hammerstrom, Donald J. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Makhmalbaf, Atefe [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Marinovici, Maria C. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2016-12-30

    Energy management in buildings is becoming more transactive. Pacific Northwest National Laboratory and the U.S. Department of Energy Building Technologies Office recently defined innovative use cases wherein market-like mechanisms are used to manage energy within buildings, between buildings, and between buildings and third-party entities, such as power utilities. A next step toward defining a set of transactive use cases in the buildings domain is to carefully diagram the corresponding business cases to capture details of transactions among all stakeholders and their economic value propositions. The principles of e3-value diagramming are applied in this report toward creating business value diagrams. These principles are extended to be consistent with Universal Modeling Language use-case diagrams. Example diagrams are presented for a subset of buildings-domain use cases that were introduced in an earlier Pacific Northwest National Laboratory report. The diagrams are intended to clearly represent an understanding of the transactions through which individual entities accumulate value in their respective use cases, and the diagrams should therefore support economic valuation studies. The report reviews some of the foundational principles of e3 value and includes authors’ insights concerning the formulation of these diagrams using Universal Modeling Language as a more systematic modeling approach.

  14. A conserved role for the ARC1 E3 ligase in Brassicaceae self-incompatibility

    Directory of Open Access Journals (Sweden)

    Daphne eGoring

    2014-05-01

    Full Text Available Ubiquitination plays essential roles in the regulation of many processes in plants including pollen rejection in self-incompatible species. In the Brassicaceae (mustard family, self-incompatibility drives the rejection of self-pollen by preventing pollen hydration following pollen contact with the stigmatic surface. Self-pollen is recognized by a ligand-receptor pair: the pollen S-locus Cysteine Rich/S-locus Protein 11 (SCR/SP11 ligand and the pistil S Receptor Kinase (SRK. Following self-pollen contact, the SCR/SP11 ligand on the pollen surface binds to SRK on the pistil surface, and the SRK-activated signaling pathway is initiated. This pathway includes the ARM Repeat Containing 1 (ARC1 protein, a member of the Plant U-box (PUB family of E3 ubiquitin ligases. ARC1 is a functional E3 ligase and is required downstream of SRK for the self-incompatibility response. This mini review highlights our recent progress in establishing ARC1’s conserved role in self-pollen rejection in Brassica and Arabidopsis species and discusses future research directions in this field.

  15. A comparative study on the distribution of non-specific esterase amongst the various constituents of retinas of some vertebrates.

    Science.gov (United States)

    Tyagi, H R; Tewari, H B

    1979-01-01

    A wide coverage of the retinae of a large number of animals (Calotes, Varanus, Naja, Athene, Passer, Streptopelia, Psittacula and Funambulus) from the point of view of the histoenzymological distribution of non-specific esterase amongst the various constituents reveals mostly identical patterns. They are as follows: 1. Outer segments - positive in all cases. 2. Outer plexiform layer - equipped with enzymatic activity in all the instances. 3. Inner nuclear layer - thin cytoplasmic rim of the neurons characterized by positive activity; the nuclei of the neurons are completely negative. 4. Inner plexiform layer - this layer is endowed with the enzymatic activity. 5. Ganglion cells - negative in all cases. 6. Nerve fibres of the layer of nerve fibres, situated adjasent to ganglion cells are positive in all the animals; in case of squirrel oligodandroglial cells present in the region have demonstrated activity of a high order. On of the high-lights of the present contribution is demarcation of the inner plexiform layer into three stratified zones, equipped with enzymatic activity in Calotes, Streptopelia, Naja and Funambulus. Such stratifications are not seen in Varanus, Passer and Psittacula. The significance of the various patterns and the equipment of the enzyme in various constituents at various locals have been discussed in relation to the metabolic functions, zone-wise and interzone-wise in visual processes of various animals.

  16. Interactions of p-Nitrobenzene Diazonium Fluoroborate and Analogs with the Active Sites of Acetylcholine-Receptor and -Esterase*

    Science.gov (United States)

    Mautner, Henry G.; Bartels, Eva

    1970-01-01

    p-Nitrobenzene diazonium fluoroborate (NDF) is a potent inhibitor of the carbamylcholine-induced depolarization of the electroplax and of acetylcholinesterase. It probably forms covalent bonds with the acetylcholine-receptor and -esterase at the active site of the proteins. Its inhibitory strength is at least the same as that of trimethylammonium diazonium fluoroborate (TDF). The p-acetoxy analog, with its weaker electron-withdrawing group, is about ten times weaker as an inhibitor than the trimethylammonium or p-nitro analogs, both of which have strong electron-withdrawing groups. After treatment of the electroplax preparation with dithiothreitol, NDF remains an irreversible receptor-inhibitor, while TDF becomes a potent reversible receptor-activator. TDF is self-inhibitory: applied before reduction, it no longer depolarizes. Although the first observations on TDF suggested that the compound labels both proteins by virtue of the steric complementary of its trimethylammonium group to a negative subsite in the proteins, the present study indicates that it is the positively charged diazonium group that reacts with the active sites of the proteins to form a covalent bond with an appropriate amino-acid residue. PMID:5272331

  17. Expression of a Petunia inflata pectin methyl esterase in Solanum tuberosum L. enhances stem elongation and modifies cation distribution.

    Science.gov (United States)

    Pilling, J; Willmitzer, L; Fisahn, J

    2000-02-01

    Transgenic potato (Solanum tuberosum L.) plants were constructed with a Petunia inflata-derived cDNA encoding a pectin methyl esterase (PME; EC 3.1.1.11) in sense orientation under the control of the cauliflower mosaic virus 35S promoter. The PME activity was elevated in leaves and tubers of the transgenic lines but slightly reduced in apical segments of stems from mature plants. Stem segments from the base of juvenile PME-overexpressing plants did not differ in PME activity from the control, whereas in apical parts PME was less active than in the wild-type. During the early stages of development stems of these transgenic plants elongated more rapidly than those of the wild-type. Further evidence that overexpression of a plant-derived PME has an impact on plant development is based on modifications of tuber yield, which was reduced in the transgenic lines. Cell walls from transgenic tubers showed significant differences in their cation-binding properties in comparison with the wild-type. In particular, cell walls displayed increased affinity for sodium and calcium, while potassium binding was constant. Furthermore, the total ion content of transgenic potatoes was modified. Indications of PME-mediated differences in the distribution of ions in transgenic plants were also obtained by monitoring relaxations of the membrane potential of roots subsequent to changes in the ionic composition of the bathing solution. However, no effects on the chemical structure of pectin from tuber cell walls could be detected.

  18. Mode of de-esterification of alkaline and acidic pectin methyl esterases at different pH conditions.

    Science.gov (United States)

    Duvetter, Thomas; Fraeye, Ilse; Sila, Daniel N; Verlent, Isabel; Smout, Chantal; Hendrickx, Marc; Van Loey, Ann

    2006-10-04

    Highly esterified citrus pectin was de-esterified at pH 4.5 and 8.0 by a fungal pectin methyl esterase (PME) that was shown to have an acidic isoelectric pH (pI) and an acidic pH optimum and by a plant PME that was characterized by an alkaline pI and an alkaline pH optimum. Interchain and intrachain de-esterification patterns were studied by digestion of the pectin products with endo-polygalacturonase and subsequent analysis using size exclusion and anion-exchange chromatography. No effect of pH was observed on the de-esterification mode of either of the two enzymes. Acidic, fungal PME converted pectin according to a multiple-chain mechanism, with a limited degree of multiple attack at the intrachain level, both at pH 4.5 and at pH 8.0. A multiple-attack mechanism, with a high degree of multiple attack, was more appropriate to describe the action mode of alkaline, plant PME, both at pH 4.5 and at pH 8.0.

  19. Textural properties of gelling system of low-methoxy pectins produced by demethoxylating reaction of pectin methyl esterase.

    Science.gov (United States)

    Kim, Y; Yoo, Y-H; Kim, K-O; Park, J-B; Yoo, S-H

    2008-06-01

    After deesterification of commercial pectins with a pectin methyl esterase (PME), their gelling properties were characterized using instrumental texture analysis. The final degree of esterification (DE) of the high- and low-methoxy pectins reached approximately 6% after the PME treatment, while deesterification of low-methoxy amidated pectin stopped at 18% DE. Furthermore, DE of high-methoxy pectin was tailored to be 40%, which is equivalent to the DE of commercial low-methoxy pectin. As a result, significant changes in molecular weight (Mw) distribution were observed in the PME-treated pectins. The texture profile analysis showed that PME modification drastically increased hardness, gumminess, and chewiness, while decreasing cohesiveness and adhesiveness of the pectin gels (P pectin gel with relatively high peak molecular weight (Mp, 3.5 x 10(5)) and low DE (6), which was produced from high-methoxy pectin, exhibited the greatest hardness, gumminess, chewiness, and resilience. The hardness of low-methoxy amidated pectin increased over 300% after PME deesterification, suggesting that the effects of amide substitution could be reinforced when DE is even lower. The partial least square regression analysis indicated that the Mw and DE of the pectin molecule are the most crucial factors for hardness, chewiness, gumminess, and resilience of gel matrix.

  20. Pectin methyl esterase treatment on high-methoxy pectin for making fruit jam with reduced sugar content.

    Science.gov (United States)

    Wang, Yuh-Tai; Lien, Ling-Lan; Chang, Ya-Chu; Wu, James Swi-Bea

    2013-01-01

    Pectin methyl esterase (PME) has been postulated to catalyse the transacylation reaction between pectin molecules. The present study aimed to prove the occurrence of this reaction. The feasibility of applying PME-catalysed transacylation between high-methoxy pectin molecules in making fruit jam with reduced sugar content was also investigated. PME treatment increased the turbidity and particle size in pectin solution and the molecular weight of pectin, while it decreased the number of methoxy ester linkages and the intensity of the CH₃ absorption peak in the Fourier transform infrared spectrum without changes in the number of total ester linkages in pectin molecules. These findings support the occurrence of PME-catalysed transacylation between pectin molecules. Higher values of hardness, gumminess and chewiness were found in a jam containing PME-treated citrus pectin (10 g L⁻¹) and sugar (350 g L⁻¹) as compared with either a jam containing untreated citrus pectin (10 g L⁻¹) and sugar (350 g L⁻¹) or strawberry jam containing pectin (10 g L⁻¹) from the fruit and sugar (650 g L⁻¹). The demand for sugar in jam making can be greatly reduced by the use of PME-treated high-methoxy pectin. Copyright © 2012 Society of Chemical Industry.

  1. The ectopic expression of a pectin methyl esterase inhibitor increases pectin methyl esterification and limits fungal diseases in wheat.

    Science.gov (United States)

    Volpi, Chiara; Janni, Michela; Lionetti, Vincenzo; Bellincampi, Daniela; Favaron, Francesco; D'Ovidio, Renato

    2011-09-01

    Cell wall pectin methyl esterification can influence plant resistance because highly methyl-esterified pectin can be less susceptible to the hydrolysis by pectic enzymes such as fungal endopolygalacturonases (PG). Pectin is secreted into the cell wall in a highly methyl-esterified form and, here, is de-methyl esterified by pectin methyl esterase (PME). The activity of PME is controlled by specific protein inhibitors called PMEI; consequently, an increased inhibition of PME by PMEI might modify the pectin methyl esterification. In order to test the possibility of improving wheat resistance by modifying the methyl esterification of pectin cell wall, we have produced durum wheat transgenic lines expressing the PMEI from Actinidia chinensis (AcPMEI). The expression of AcPMEI endows wheat with a reduced endogenous PME activity, and transgenic lines expressing a high level of the inhibitor showed a significant increase in the degree of methyl esterification. These lines showed a significant reduction of disease symptoms caused by the fungal pathogens Bipolaris sorokiniana or Fusarium graminearum. This increased resistance was related to the impaired ability of these fungal pathogens to grow on methyl-esterified pectin and to a reduced activity of the fungal PG to hydrolyze methyl-esterified pectin. In addition to their importance for wheat improvement, these results highlight the primary role of pectin despite its low content in the wheat cell wall.

  2. Partial demethylation of oligogalacturonides by pectin methyl esterase 1 is required for eliciting defence responses in wild strawberry (Fragaria vesca).

    Science.gov (United States)

    Osorio, Sonia; Castillejo, Cristina; Quesada, Miguel A; Medina-Escobar, Nieves; Brownsey, Geoff J; Suau, Rafael; Heredia, Antonio; Botella, Miguel A; Valpuesta, Victoriano

    2008-04-01

    In addition to the role of the cell wall as a physical barrier against pathogens, some of its constituents, such as pectin-derived oligogalacturonides (OGA), are essential components for elicitation of defence responses. To investigate how modifications of pectin alter defence responses, we expressed the fruit-specific Fragaria x ananassa pectin methyl esterase FaPE1 in the wild strawberry Fragaria vesca. Pectin from transgenic ripe fruits differed from the wild-type with regard to the degree and pattern of methyl esterification, as well as the average size of pectin polymers. Purified oligogalacturonides from the transgenic fruits showed a reduced degree of esterification compared to oligogalacturonides from wild-type fruits. This reduced esterification is necessary to elicit defence responses in strawberry. The transgenic F. vesca lines had constitutively activated pathogen defence responses, resulting in higher resistance to the necrotropic fungus Botrytis cinerea. Further studies in F. vesca and Nicotiana benthamiana leaves showed that the elicitation capacity of the oligogalacturonides is more specific than previously envisaged.

  3. Improved stability and catalytic activity of graphene oxide/chitosan hybrid beads loaded with porcine liver esterase.

    Science.gov (United States)

    Sunderrajan, Shruthi; Miranda, Lima Rose; Pennathur, Gautam

    2018-04-21

    Graphene oxide/chitosan and reduced graphene oxide/chitosan (GO/CS and RGO/CS) beads were prepared by precipitation with NaOH. Porcine liver esterase was immobilized on these beads to give GO/CS/E and RGO/CS/E beads. The optimum conditions for the maximum activity of RGO/CS/E beads were pH 8 and 50°C. The stability of the enzyme immobilized on GO/CS/E and RGO/CS/E was high in the pH range of 5-8. The GO/CS/E beads showed superior stability compared to that of the free enzyme and CS/E beads between 20 and 50°C. Kinetic analysis showed that GO/CS/E was a better catalyst than the RGO/CS/E beads with a lower K m value of 0.9 mM. The hybrid beads also retained more than 95% activity after 10 consecutive cycles. The GO/CS/E and RGO/CS/E beads retained 84% and 87% activity after 40 days at 4°C. The GO/CS/E beads were used for the successful hydrolysis of methyl 4-hydroxy benzoate.

  4. Probiotic Ferulic Acid Esterase Active Lactobacillus fermentum NCIMB 5221 APA Microcapsules for Oral Delivery: Preparation and in Vitro Characterization

    Directory of Open Access Journals (Sweden)

    Catherine Tomaro-Duchesneau

    2012-02-01

    Full Text Available Probiotics possess potential therapeutic and preventative effects for various diseases and metabolic disorders. One important limitation for the oral delivery of probiotics is the harsh conditions of the upper gastrointestinal tract (GIT which challenge bacterial viability and activity. One proposed method to surpass this obstacle is the use of microencapsulation to improve the delivery of bacterial cells to the lower GIT. The aim of this study is to use alginate-poly-L-lysine-alginate (APA microcapsules to encapsulate Lactobacillus fermentum NCIMB 5221 and characterize its enzymatic activity and viability through a simulated GIT. This specific strain, in previous research, was characterized for its inherent ferulic acid esterase (FAE activity which could prove beneficial in the development of a therapeutic for the treatment and prevention of cancers and metabolic disorders. Our findings demonstrate that the APA microcapsule does not slow the mass transfer of substrate into and that of the FA product out of the microcapsule, while also not impairing bacterial cell viability. The use of simulated gastrointestinal conditions led to a significant 2.5 log difference in viability between the free (1.10 × 104 ± 1.00 × 103 cfu/mL and the microencapsulated (5.50 × 106 ± 1.00 × 105 cfu/mL L. fermentum NCIMB 5221 following exposure. The work presented here suggests that APA microencapsulation can be used as an effective oral delivery method for L. fermentum NCIMB 5221, a FAE-active probiotic strain.

  5. Probiotic Ferulic Acid Esterase Active Lactobacillus fermentum NCIMB 5221 APA Microcapsules for Oral Delivery: Preparation and in Vitro Characterization

    Science.gov (United States)

    Tomaro-Duchesneau, Catherine; Saha, Shyamali; Malhotra, Meenakshi; Coussa-Charley, Michael; Kahouli, Imen; Jones, Mitchell L.; Labbé, Alain; Prakash, Satya

    2012-01-01

    Probiotics possess potential therapeutic and preventative effects for various diseases and metabolic disorders. One important limitation for the oral delivery of probiotics is the harsh conditions of the upper gastrointestinal tract (GIT) which challenge bacterial viability and activity. One proposed method to surpass this obstacle is the use of microencapsulation to improve the delivery of bacterial cells to the lower GIT. The aim of this study is to use alginate-poly-L-lysine-alginate (APA) microcapsules to encapsulate Lactobacillus fermentum NCIMB 5221 and characterize its enzymatic activity and viability through a simulated GIT. This specific strain, in previous research, was characterized for its inherent ferulic acid esterase (FAE) activity which could prove beneficial in the development of a therapeutic for the treatment and prevention of cancers and metabolic disorders. Our findings demonstrate that the APA microcapsule does not slow the mass transfer of substrate into and that of the FA product out of the microcapsule, while also not impairing bacterial cell viability. The use of simulated gastrointestinal conditions led to a significant 2.5 log difference in viability between the free (1.10 × 104 ± 1.00 × 103 cfu/mL) and the microencapsulated (5.50 × 106 ± 1.00 × 105 cfu/mL) L. fermentum NCIMB 5221 following exposure. The work presented here suggests that APA microencapsulation can be used as an effective oral delivery method for L. fermentum NCIMB 5221, a FAE-active probiotic strain. PMID:24288090

  6. Leukocyte esterase reagent strip as a bedside tool to detect peritonitis in patients undergoing acute peritoneal dialysis

    Directory of Open Access Journals (Sweden)

    Vinay Rathore

    2017-01-01

    Full Text Available Peritonitis is a common and life-threatening complication of acute peritoneal dialysis (PD. Diagnosis requires the presence of clinical signs of peritonitis which are nonspecific and laboratory investigations [total leukocyte count (TLC, Gram-stain, and culture of PD effluent fluid] which are time-consuming and not available at the bedside. In this study, we evaluated the use of leukocyte esterase reagent strip (LERS as a bedside test to diagnose peritonitis in patients undergoing acute PD. Patients who underwent acute PD were monitored for signs and symptoms of peritonitis. PD effluent fluid analysis included TLC, absolute neutrophil count, Gram-stain, and culture for the diagnosis of peritonitis. LERS (Multistix 10SG was simultaneously dipped in PD effluent fluid and read at two minutes. Reading of + was considered as indicative of peritonitis. Twenty-one out of 166 (12.6% patients undergoing acute PD developed peritonitis. LERS detected peritonitis in 20 patients. The sensitivity, specificity, positive predictive value, and negative predictive value (NPV of LERS were 95.2%, 95.2%, 74.1%, and 99.3%, respectively. LERS has very high sensitivity and NPV and can be used as a rapid bedside tool to exclude peritonitis in patients undergoing acute PD.

  7. Selective Activation of N,N'-Diacyl Rhodamine Pro-fluorophores Paired with Releasing Enzyme, Porcine Liver Esterase (PLE).

    Science.gov (United States)

    Abney, Kristopher K; Ramos-Hunter, Susan J; Romaine, Ian M; Godwin, J Shawn; Sulikowski, Gary A; Weaver, Charles David

    2018-04-21

    This study reports the synthesis and testing of a family of rhodamine pro-fluorophores and an enzyme capable of converting pro-fluorophores to Rhodamine 110. We prepared a library of simple N,N'-diacyl rhodamines and investigated Porcine Liver Esterase (PLE) as an enzyme to activate rhodamine-based pro-fluorophores. A PLE-expressing cell line generated an increase in fluorescence rapidly upon pro-fluorophore addition demonstrating the rhodamine pro-fluorophores are readily taken up and fluorescent upon PLE-mediated release. Rhodamine pro-fluorophore amides trifluoroacetamide (TFAm) and proponamide (PAm) appeared to be the best substrates using a cell-based assay using PLE expressing HEK293. Our pro-fluorophore series showed diffusion into live cells and resisted endogenous hydrolysis. The use of our engineered cell line containing the exogenous enzyme PLE demonstrated the rigorousness of amide masking when compared to cells not containing PLE. This simple and selective pro-fluorophore rhodamine pair with PLE offers the potential to be used in vitro and in vivo fluorescence based assays. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Probiotic Ferulic Acid Esterase Active Lactobacillus fermentum NCIMB 5221 APA Microcapsules for Oral Delivery: Preparation and in Vitro Characterization.

    Science.gov (United States)

    Tomaro-Duchesneau, Catherine; Saha, Shyamali; Malhotra, Meenakshi; Coussa-Charley, Michael; Kahouli, Imen; Jones, Mitchell L; Labbé, Alain; Prakash, Satya

    2012-02-16

    Probiotics possess potential therapeutic and preventative effects for various diseases and metabolic disorders. One important limitation for the oral delivery of probiotics is the harsh conditions of the upper gastrointestinal tract (GIT) which challenge bacterial viability and activity. One proposed method to surpass this obstacle is the use of microencapsulation to improve the delivery of bacterial cells to the lower GIT. The aim of this study is to use alginate-poly-L-lysine-alginate (APA) microcapsules to encapsulate Lactobacillus fermentum NCIMB 5221 and characterize its enzymatic activity and viability through a simulated GIT. This specific strain, in previous research, was characterized for its inherent ferulic acid esterase (FAE) activity which could prove beneficial in the development of a therapeutic for the treatment and prevention of cancers and metabolic disorders. Our findings demonstrate that the APA microcapsule does not slow the mass transfer of substrate into and that of the FA product out of the microcapsule, while also not impairing bacterial cell viability. The use of simulated gastrointestinal conditions led to a significant 2.5 log difference in viability between the free (1.10 × 104 ± 1.00 × 103 cfu/mL) and the microencapsulated (5.50 × 106 ± 1.00 × 105 cfu/mL) L. fermentum NCIMB 5221 following exposure. The work presented here suggests that APA microencapsulation can be used as an effective oral delivery method for L. fermentum NCIMB 5221, a FAE-active probiotic strain.

  9. Effects of juvenile hormone (JH) analog insecticides on larval development and JH esterase activity in two spodopterans.

    Science.gov (United States)

    El-Sheikh, El-Sayed A; Kamita, Shizuo G; Hammock, Bruce D

    2016-03-01

    Juvenile hormone analog (JHA) insecticides are biological and structural mimics of JH, a key insect developmental hormone. Toxic and anti-developmental effects of the JHA insecticides methoprene, fenoxycarb, and pyriproxyfen were investigated on the larval and pupal stages of Spodoptera littoralis and Spodoptera frugiperda. Bioassays showed that fenoxycarb has the highest toxicity and fastest speed of kill in 2nd instar S. littoralis. All three JHAs affected the development of 6th instar (i.e., final instar) and pupal S. frugiperda. JH esterase (JHE) is a critical enzyme that helps to regulate JH levels during insect development. JHE activity in the last instar S. littoralis and S. frugiperda was 11 and 23 nmol min(-1) ml(-1) hemolymph, respectively. Methoprene and pyriproxyfen showed poor inhibition of JHE activity from these insects, whereas fenoxycarb showed stronger inhibition. The inhibitory activity of fenoxycarb, however, was more than 1000-fold lower than that of OTFP, a highly potent inhibitor of JHEs. Surprisingly, topical application of methoprene, fenoxycarb or pyriproxyfen on 6th instars of S. littoralis and S. frugiperda prevented the dramatic reduction in JHE activity that was found in control insects. Our findings suggest that JHAs may function as JH agonists that play a disruptive role or a hormonal replacement role in S. littoralis and S. frugiperda. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Genes and Gene Therapy

    Science.gov (United States)

    ... correctly, a child can have a genetic disorder. Gene therapy is an experimental technique that uses genes to ... or prevent disease. The most common form of gene therapy involves inserting a normal gene to replace an ...

  11. Time-of-day- and light-dependent expression of ubiquitin protein ligase E3 component N-recognin 4 (UBR4 in the suprachiasmatic nucleus circadian clock.

    Directory of Open Access Journals (Sweden)

    Harrod H Ling

    Full Text Available Circadian rhythms of behavior and physiology are driven by the biological clock that operates endogenously but can also be entrained to the light-dark cycle of the environment. In mammals, the master circadian pacemaker is located in the suprachiasmatic nucleus (SCN, which is composed of individual cellular oscillators that are driven by a set of core clock genes interacting in transcriptional/translational feedback loops. Light signals can trigger molecular events in the SCN that ultimately impact on the phase of expression of core clock genes to reset the master pacemaker. While transcriptional regulation has received much attention in the field of circadian biology in the past, other mechanisms including targeted protein degradation likely contribute to the clock timing and entrainment process. In the present study, proteome-wide screens of the murine SCN led to the identification of ubiquitin protein ligase E3 component N-recognin 4 (UBR4, a novel E3 ubiquitin ligase component of the N-end rule pathway, as a time-of-day-dependent and light-inducible protein. The spatial and temporal expression pattern of UBR4 in the SCN was subsequently characterized by immunofluorescence microscopy. UBR4 is expressed across the entire rostrocaudal extent of the SCN in a time-of-day-dependent fashion. UBR4 is localized exclusively to arginine vasopressin (AVP-expressing neurons of the SCN shell. Upon photic stimulation in the early subjective night, the number of UBR4-expressing cells within the SCN increases. This study is the first to identify a novel E3 ubiquitin ligase component, UBR4, in the murine SCN and to implicate the N-end rule degradation pathway as a potential player in regulating core clock mechanisms and photic entrainment.

  12. Differential effects of amlodipine and atorvastatin treatment and their combination on atherosclerosis in ApoE*3-Leiden transgenic mice

    NARCIS (Netherlands)

    Delsing, D.J.; Jukema, J.W.; van de Wiel, M.A.; Emeis, J.; van der Laarse, A.; Havekes, L.M.; Princen, H.M.G.

    2003-01-01

    This study was designed to investigate the potential antiatherosclerotic effects of the calcium antagonist amlodipine as compared with the HMG-CoA reductase inhibitor atorvastatin and the combination of both in ApoE*3-Leiden transgenic mice. Four groups of 15 ApoE*3-Leiden mice were put on a

  13. Structural analysis of lunar subsurface with Chang'E-3 lunar penetrating radar

    Science.gov (United States)

    Lai, Jialong; Xu, Yi; Zhang, Xiaoping; Tang, Zesheng

    2016-01-01

    Geological structure of the subsurface of the Moon provides valuable information on lunar evolution. Recently, Chang'E-3 has utilized lunar penetrating radar (LPR), which is equipped on the lunar rover named as Yutu, to detect the lunar geological structure in Northern Imbrium (44.1260N, 19.5014W) for the first time. As an in situ detector, Chang'E-3 LPR has relative higher horizontal and vertical resolution and less clutter impact compared to spaceborne radars and earth-based radars. In this work, we analyze the LPR data at 500 MHz transmission frequency to obtain the shallow subsurface structure of the landing area of Chang'E-3 in Mare Imbrium. Filter method and amplitude recovery algorithms are utilized to alleviate the adverse effects of environment and system noises and compensate the amplitude losses during signal propagation. Based on the processed radar image, we observe numerous diffraction hyperbolae, which may be caused by discrete reflectors beneath the lunar surface. Hyperbolae fitting method is utilized to reverse the average dielectric constant to certain depth (ε bar). Overall, the estimated ε bar increases with the depth and ε bar could be classified into three categories. Average ε bar of each category is 2.47, 3.40 and 6.16, respectively. Because of the large gap between the values of ε bar of neighboring categories, we speculate a three-layered structure of the shallow surface of LPR exploration region. One possible geological picture of the speculated three-layered structure is presented as follows. The top layer is weathered layer of ejecta blanket with its average thickness and bound on error is 0.95±0.02 m. The second layer is the ejecta blanket of the nearby impact crater, and the corresponding average thickness is about 2.30±0.07 m, which is in good agreement with the two primary models of ejecta blanket thickness as a function of distance from the crater center. The third layer is regarded as a mixture of stones and soil. The

  14. Structural Analysis of Lunar Subsurface with Chang'E 3 Lunar Penetrating Radar

    Science.gov (United States)

    Xu, Yi; Lai, Jialong; Tang, Zesheng

    2015-04-01

    Geological structure of the subsurface of the Moon provides valuable information for our understanding of lunar evolution. Recently, Chang'E 3 has utilized lunar penetrating radar (LPR), which is equipped on the lunar rover named as Yutu, to detect the lunar geological structure in Northern Imbrium (44.1260N, 19.5014W) for the first time. As an in-situ detector, Chang'E 3 LPR has higher horizontal and vertical resolution and less clutter impact compared to spaceborne radars such as Chandrayaan-1 and Kaguya. In this work, we analyze the LPR data at 500 MHz transmission frequency to obtain the shallow subsurface structure of the landing area of Chang'E 3 in Mare Imbrium. First, filter method and amplitude recover algorithms are introduced for data processing to alleviate the adverse effects of environment and system noises and compensate the amplitude losses during signal propagation. Next, based on the processed LPR data, we present the methods to determine the interfaces between layers. A three-layered structure of the shallow surface of the Moon has been observed. The corresponding real part of relative dielectric constant is inverted with deconvolution method. The average dielectric constants of the surface, second and third layer is 2.8, 3.2 and 3.6, respectively. The phenomenon that the average dielectric constant increases with the depth is consistent with prior art. With the obtained dielectric constants, the thickness of each layer can be calculated. One possible geological picture of the observed three-layered structure is presented as follows. The top layer is lunar regolith with its thickness ranging from 0.59 m to 0.9 m. The second layer is the ejecta blanket of the nearby impact crater, and the corresponding thickness is between 3.6m to 3.9m, which is in good agreement with the model of ejecta blanket thickness (height) as a function of distance from the crater center proposed by Melosh in 1989. The third layer is regarded as early lunar regolith with 4

  15. Global economics/energy/environmental (E3) modeling of long-term nuclear energy futures

    International Nuclear Information System (INIS)

    Krakowski, R.A.; Davidson, J.W.; Bathke, C.G.; Arthur, E.D.; Wagner, R.L. Jr.

    1997-01-01

    A global energy, economics, environment (E 3 ) model has been adopted and modified with a simplified, but comprehensive and multi-regional, nuclear energy module. Using this model, consistent nuclear energy scenarios are constructed. A spectrum of future is examined at two levels in a hierarchy of scenario attributes in which drivers are either external or internal to nuclear energy. Impacts of a range of nuclear fuel-cycle scenarios are reflected back to the higher-level scenario attributes. An emphasis is placed on nuclear materials inventories (in magnitude, location, and form) and their contribution to the long-term sustainability of nuclear energy and the future competitiveness of both conventional and advanced nuclear reactors

  16. Design and Experimental Verification of Chang'E-3 Moon-night Survival Device for APXS

    Science.gov (United States)

    Deng-yi, Chen; Jian, Wu; Yi-ming, Hu; Jin, Chang; Yi-zhong, Gong; Ming-sheng, Cai; Huan-yu, Wang; Jia-yu, Zhang; Xing-zhu, Cui; Jin-zhou, Wang

    2016-07-01

    The Active Particle X-ray Spectrometer (APXS) is one of the 4 scientific payloads of Chang'E-3 (CE-3) Lunar Rover, of which the scientific object is to identify the elements of lunar soil and rock samples by a carried radioactive source to trigger and detect the characteristic X-ray from them. According to the extreme temperature environment of the APXS and under the restriction of limited resources, this paper presents the design and analysis of the moon-night survival device RHU (radioisotope heating unit) for the APXS, and describes the corresponding environmental tests on its structure dynamics and moon-night survival. Finally, its reinstallation on the launch tower and the preliminary result of its on-orbit operation are introduced.

  17. Data processing and initial results of Chang'e-3 lunar penetrating radar

    Science.gov (United States)

    Su, Yan; Fang, Guang-You; Feng, Jian-Qing; Xing, Shu-Guo; Ji, Yi-Cai; Zhou, Bin; Gao, Yun-Ze; Li, Han; Dai, Shun; Xiao, Yuan; Li, Chun-Lai

    2014-12-01

    To improve our understanding of the formation and evolution of the Moon, one of the payloads onboard the Chang'e-3 (CE-3) rover is Lunar Penetrating Radar (LPR). This investigation is the first attempt to explore the lunar subsurface structure by using ground penetrating radar with high resolution. We have probed the subsurface to a depth of several hundred meters using LPR. In-orbit testing, data processing and the preliminary results are presented. These observations have revealed the configuration of regolith where the thickness of regolith varies from about 4 m to 6 m. In addition, one layer of lunar rock, which is about 330 m deep and might have been accumulated during the depositional hiatus of mare basalts, was detected.

  18. Data processing and initial results of Chang'e-3 lunar penetrating radar

    International Nuclear Information System (INIS)

    Su Yan; Feng Jian-Qing; Xing Shu-Guo; Li Han; Dai Shun; Xiao Yuan; Li Chun-Lai; Fang Guang-You; Ji Yi-Cai; Zhou Bin; Gao Yun-Ze

    2014-01-01

    To improve our understanding of the formation and evolution of the Moon, one of the payloads onboard the Chang'e-3 (CE-3) rover is Lunar Penetrating Radar (LPR). This investigation is the first attempt to explore the lunar subsurface structure by using ground penetrating radar with high resolution. We have probed the subsurface to a depth of several hundred meters using LPR. In-orbit testing, data processing and the preliminary results are presented. These observations have revealed the configuration of regolith where the thickness of regolith varies from about 4 m to 6 m. In addition, one layer of lunar rock, which is about 330 m deep and might have been accumulated during the depositional hiatus of mare basalts, was detected

  19. A young multilayered terrane of the northern Mare Imbrium revealed by Chang’E-3 mission

    Science.gov (United States)

    Xiao, Long; Zhu, Peimin; Fang, Guangyou; Xiao, Zhiyong; Zou, Yongliao; Zhao, Jiannan; Zhao, Na; Yuan, Yuefeng; Qiao, Le; Zhang, Xiaoping; Zhang, Hao; Wang, Jiang; Huang, Jun; Huang, Qian; He, Qi; Zhou, Bin; Ji, Yicai; Zhang, Qunying; Shen, Shaoxiang; Li, Yuxi; Gao, Yunze

    2015-03-01

    China’s Chang’E-3 (CE-3) spacecraft touched down on the northern Mare Imbrium of the lunar nearside (340.49°E, 44.12°N), a region not directly sampled before. We report preliminary results with data from the CE-3 lander descent camera and from the Yutu rover’s camera and penetrating radar. After the landing at a young 450-meter crater rim, the Yutu rover drove 114 meters on the ejecta blanket and photographed the rough surface and the excavated boulders. The boulder contains a substantial amount of crystals, which are most likely plagioclase and/or other mafic silicate mineral aggregates similar to terrestrial dolerite. The Lunar Penetrating Radar detection and integrated geological interpretation have identified more than nine subsurface layers, suggesting that this region has experienced complex geological processes since the Imbrian and is compositionally distinct from the Apollo and Luna landing sites.

  20. The preliminary processing and analysis of LPR Channel-2B data from Chang'E-3

    Science.gov (United States)

    Zhao, Na; Zhu, PeiMin; Yang, KeSi; Yuan, YueFeng; Guo, ShiLi

    2014-12-01

    The Lunar Penetrating Radar (LPR) carried by Chang'E-3 has imaged the shallow subsurface of the landing site at the northern Mare Imbrium. The antenna B of the Channel-2 onboard the LPR (LPR Channel-2B) has collected more than 2000 traces of usable raw data. Because of the low resolution and noise of the raw data, only a few shallow geological structures are visible. To improve the resolution and the signal-to-noise ratio of the LPR data, we processed the LPR data including amplitude compensation, filtering, and deconvolution processes. The processing results reveal that the data processing in this study not only improves the signal-to-noise ratio of the LPR Channel-2B data but also makes the geological structures vivid. The processing results will lay the foundation for the subsequent geological interpretation and physical property inversion of lunar materials.

  1. Regolith stratigraphy at the Chang'E-3 landing site as seen by lunar penetrating radar

    Science.gov (United States)

    Fa, Wenzhe; Zhu, Meng-Hua; Liu, Tiantian; Plescia, Jeffrey B.

    2015-12-01

    The Chang'E-3 lunar penetrating radar (LPR) observations at 500 MHz reveal four major stratigraphic zones from the surface to a depth of ~20 m along the survey line: a layered reworked zone (<1 m), an ejecta layer (~2-6 m), a paleoregolith layer (~4-11 m), and the underlying mare basalts. The reworked zone has two to five distinct layers and consists of surface regolith. The paleoregolith buried by the ejecta from a 500 m crater is relatively homogenous and contains only a few rocks. Population of buried rocks increases with depth to ~2 m at first, and then decreases with depth, representing a balance between initial deposition of the ejecta and later turnover of the regolith. Combining with the surface age, the LPR observations indicate a mean accumulation rate of about 5-10 m/Gyr for the surface regolith, which is at least 4-8 times larger than previous estimation.

  2. A young multilayered terrane of the northern Mare Imbrium revealed by Chang'E-3 mission.

    Science.gov (United States)

    Xiao, Long; Zhu, Peimin; Fang, Guangyou; Xiao, Zhiyong; Zou, Yongliao; Zhao, Jiannan; Zhao, Na; Yuan, Yuefeng; Qiao, Le; Zhang, Xiaoping; Zhang, Hao; Wang, Jiang; Huang, Jun; Huang, Qian; He, Qi; Zhou, Bin; Ji, Yicai; Zhang, Qunying; Shen, Shaoxiang; Li, Yuxi; Gao, Yunze

    2015-03-13

    China's Chang'E-3 (CE-3) spacecraft touched down on the northern Mare Imbrium of the lunar nearside (340.49°E, 44.12°N), a region not directly sampled before. We report preliminary results with data from the CE-3 lander descent camera and from the Yutu rover's camera and penetrating radar. After the landing at a young 450-meter crater rim, the Yutu rover drove 114 meters on the ejecta blanket and photographed the rough surface and the excavated boulders. The boulder contains a substantial amount of crystals, which are most likely plagioclase and/or other mafic silicate mineral aggregates similar to terrestrial dolerite. The Lunar Penetrating Radar detection and integrated geological interpretation have identified more than nine subsurface layers, suggesting that this region has experienced complex geological processes since the Imbrian and is compositionally distinct from the Apollo and Luna landing sites. Copyright © 2015, American Association for the Advancement of Science.

  3. Structure and catalytic activation of the TRIM23 RING E3 ubiquitin ligase: DAWIDZIAK et al.

    Energy Technology Data Exchange (ETDEWEB)

    Dawidziak, Daria M. [Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville Virginia; Sanchez, Jacint G. [Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville Virginia; Wagner, Jonathan M. [Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville Virginia; Ganser-Pornillos, Barbie K. [Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville Virginia; Pornillos, Owen [Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville Virginia

    2017-07-24

    Tripartite motif (TRIM) proteins comprise a large family of RING-type ubiquitin E3 ligases that regulate important biological processes. An emerging general model is that TRIMs form elongated antiparallel coiled-coil dimers that prevent interaction of the two attendant RING domains. The RING domains themselves bind E2 conjugating enzymes as dimers, implying that an active TRIM ligase requires higher-order oligomerization of the basal coiled-coil dimers. Here, we report crystal structures of the TRIM23 RING domain in isolation and in complex with an E2–ubiquitin conjugate. Our results indicate that TRIM23 enzymatic activity requires RING dimerization, consistent with the general model of TRIM activation.

  4. RING E3 ligases: key regulatory elements are involved in abiotic stress responses in plants.

    Science.gov (United States)

    Cho, Seok Keun; Ryu, Moon Young; Kim, Jong Hum; Hong, Jeong Soo; Oh, Tae Rin; Kim, Woo Taek; Yang, Seong Wook

    2017-08-01

    Plants are constantly exposed to a variety of abiotic stresses, such as drought, heat, cold, flood, and salinity. To survive under such unfavorable conditions, plants have evolutionarily developed their own resistant-mechanisms. For several decades, many studies have clarified specific stress response pathways of plants through various molecular and genetic studies. In particular, it was recently discovered that ubiquitin proteasome system (UPS), a regulatory mechanism for protein turn over, is greatly involved in the stress responsive pathways. In the UPS, many E3 ligases play key roles in recognizing and tethering poly-ubiquitins on target proteins for subsequent degradation by the 26S proteasome. Here we discuss the roles of RING ligases that have been defined in related to abiotic stress responses in plants. [BMB Reports 2017; 50(8): 393-400].

  5. Degradation of human Lipin-1 by BTRC E3 ubiquitin ligase.

    Science.gov (United States)

    Ishimoto, Kenji; Hayase, Ayaka; Kumagai, Fumiko; Kawai, Megumi; Okuno, Hiroko; Hino, Nobumasa; Okada, Yoshiaki; Kawamura, Takeshi; Tanaka, Toshiya; Hamakubo, Takao; Sakai, Juro; Kodama, Tatsuhiko; Tachibana, Keisuke; Doi, Takefumi

    2017-06-17

    Lipin-1 has dual functions in the regulation of lipid and energy metabolism according to its subcellular localization, which is tightly controlled. However, it is unclear how Lipin-1 degradation is regulated. Here, we demonstrate that Lipin-1 is degraded through its DSGXXS motif. We show that Lipin-1 interacts with either of two E3 ubiquitin ligases, BTRC or FBXW11, and that this interaction is DSGXXS-dependent and mediates the attachment of polyubiquitin chains. Further, we demonstrate that degradation of Lipin-1 is regulated by BTRC in the cytoplasm and on membranes. These novel insights into the regulation of human Lipin-1 stability will be useful in planning further studies to elucidate its metabolic processes. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Ret Finger Protein: An E3 Ubiquitin Ligase Juxtaposed to the XY Body in Meiosis

    Directory of Open Access Journals (Sweden)

    Isabelle Gillot

    2009-01-01

    Full Text Available During prophase I of male meiosis, the sex chromosomes form a compact structure called XY body that associates with the nuclear membrane of pachytene spermatocytes. Ret Finger Protein is a transcriptional repressor, able to interact with both nuclear matrix-associated proteins and double-stranded DNA. We report the precise and unique localization of Ret Finger Protein in pachytene spermatocytes, in which Ret Finger Protein takes place of lamin B1, between the XY body and the inner nuclear membrane. This localization of Ret Finger Protein does not seem to be associated with O-glycosylation or sumoylation. In addition, we demonstrate that Ret Finger Protein contains an E3 ubiquitin ligase activity. These observations lead to an attractive hypothesis in which Ret Finger Protein would be involved in the positioning and the attachment of XY body to the nuclear lamina of pachytene spermatocytes.

  7. Spectroscopy of 215Ra: the shell model and enhanced E3 transitions

    International Nuclear Information System (INIS)

    Stuchbery, A.E.; Dracoulis, G.D.; Kibedi, T.; Fabricius, B.; Lane, G.J.; Poletti, A.R.; Baxter, A.M.

    1998-01-01

    Excited states in the N=127 nucleus 215 Ra have been studied using γ-ray and electron spectroscopy following reactions of 13 C on 206 Pb targets. Levels were identified up to spins of ∝61/2 ℎ and excitation energies of ∝6 MeV. Enhanced octupole transitions are a feature of the level scheme. Lifetimes and magnetic moments were measured for several isomeric levels. The level scheme, transition rates and magnetic moments are compared with empirical shell model calculations and multiparticle octupole-coupled shell model calculations. In general, the experimental data are well described, but in comparison with its success in describing enhanced E3 transitions between related states in the radon isotopes, some limitations of the multiparticle octupole-coupling approach are revealed in 215 Ra. (orig.)

  8. Merkel cell polyomavirus small T antigen induces genome instability by E3 ubiquitin ligase targeting.

    Science.gov (United States)

    Kwun, H J; Wendzicki, J A; Shuda, Y; Moore, P S; Chang, Y

    2017-12-07

    The formation of a bipolar mitotic spindle is an essential process for the equal segregation of duplicated DNA into two daughter cells during mitosis. As a result of deregulated cellular signaling pathways, cancer cells often suffer a loss of genome integrity that might etiologically contribute to carcinogenesis. Merkel cell polyomavirus (MCV) small T (sT) oncoprotein induces centrosome overduplication, aneuploidy, chromosome breakage and the formation of micronuclei by targeting cellular ligases through a sT domain that also inhibits MCV large T oncoprotein turnover. These results provide important insight as to how centrosome number and chromosomal stability can be affected by the E3 ligase targeting capacity of viral oncoproteins such as MCV sT, which may contribute to Merkel cell carcinogenesis.

  9. [Near infrared distance sensing method for Chang'e-3 alpha particle X-ray spectrometer].

    Science.gov (United States)

    Liang, Xiao-Hua; Wu, Ming-Ye; Wang, Huan-Yu; Peng, Wen-Xi; Zhang, Cheng-Mo; Cui, Xing-Zhu; Wang, Jin-Zhou; Zhang, Jia-Yu; Yang, Jia-Wei; Fan, Rui-Rui; Gao, Min; Liu, Ya-Qing; Zhang, Fei; Dong, Yi-Fan; Guo, Dong-Ya

    2013-05-01

    Alpha particle X-ray spectrometer (APXS) is one of the payloads of Chang'E-3 lunar rover, the scientific objective of which is in-situ observation and off-line analysis of lunar regolith and rock. Distance measurement is one of the important functions for APXS to perform effective detection on the moon. The present paper will first give a brief introduction to APXS, and then analyze the specific requirements and constraints to realize distance measurement, at last present a new near infrared distance sensing algorithm by using the inflection point of response curve. The theoretical analysis and the experiment results verify the feasibility of this algorithm. Although the theoretical analysis shows that this method is not sensitive to the operating temperature and reflectance of the lunar surface, the solar infrared radiant intensity may make photosensor saturation. The solutions are reducing the gain of device and avoiding direct exposure to sun light.

  10. Socio-economic factors associated with a healthy diet: results from the E3N study.

    Science.gov (United States)

    Affret, Aurélie; Severi, Gianluca; Dow, Courtney; Rey, Grégoire; Delpierre, Cyrille; Boutron-Ruault, Marie-Christine; Clavel-Chapelon, Françoise; Fagherazzi, Guy

    2017-06-01

    To identify individual and contextual socio-economic factors associated with a healthy diet. Dietary data from a large cohort study were used to derive two mutually exclusive dietary patterns through a latent class analysis. Associations between dietary patterns and socio-economic factors were studied with logistic regression. E3N, a French prospective cohort study composed of women recruited from a national health insurance plan covering people working in the national education system. E3N participants (n 73 031) with dietary and socio-economic data available. The 'Healthy' pattern was characterized by a large consumption of fruits and vegetables and the 'Less Healthy' pattern by a large consumption of pizza and processed meat. When all socio-economic factors were analysed together, all of the individual factors considered were associated with a healthy diet (e.g. women with three or more children were less likely to follow a healthy diet v. women with no children, OR (95 % CI): 0·70 (0·66, 0·75)) while the contextual factors associated with a healthy diet included the size of the agglomeration of residence and the area of birth and residence (e.g. women living in the West of France were less likely to follow a healthy diet v. those living in the South of France: 0·78 (0·72, 0·83)). We demonstrated that individual and contextual factors are both associated with diet. Rather than focusing only on individual factors, we recommend future studies or public health and nutritional strategies on diet to consider both types of factors.

  11. Chang'e 3 and Jade Rabbit's: observations and the landing zone

    Science.gov (United States)

    Ping, Jinsong

    Chang’E-3 was launched and landed on the near side of the Moon in December 2013. It is realizing the 2nd phase of Chinese lunar scientific exploration projects. Together with the various in-situ optical observations around the landing sites, the mission carried 4 kinds of radio science experiments, cover the various lunar scientific disciplines as well as lunar surface radio astronomy studies. The key payloads onboard the lander and rover include the near ultraviolet telescope, extreme ultraviolet cameras, ground penetrating radar, very low frequency radio spectrum analyzer, which have not been used in earlier lunar landing missions. Optical spectrometer, Alpha Paticle X-ray spectrometer and Gama Ray spectrometer is also used. The mission is using extreme ultraviolet camera to observe the sun activity and geomagnetic disturbances on geo-space plasma layer of extreme ultraviolet radiation, studying space weather in the plasma layer role in the process; the mission also carries the first time lunar base optical astronomical observations. Most importantly, the topography, landforms and geological structure has been explored in detail. Additionally, the very precise Earth-Moon radio phase ranging technique was firstly tested and realized in this mission. It may increase the study of lunar dyanmics together with LLR technique. Similar to Luna-Glob landers, together with the VLBI radio beacons, the radio transponders are also set on the Chang’E-3. Transponder will receive the uplink X band radio wave transmitted from the two newly constructed Chinese deep space stations, where the high quality hydrogen maser atomic clocks have been used as local time and frequency standard. Radio science receivers have been developed by updating the multi-channel open loop Doppler receiver developed for VLBI and Doppler tracking in Yinghuo-1 and Phobos-Glob Martian missions. This experiment will improve the study of lunar dynamics, by means of measuring the lunar physical liberations

  12. The interactions in K{sub μ3} and K{sub e3} decay

    Energy Technology Data Exchange (ETDEWEB)

    McDowell, S. W. [Brazilian Centre of Research in Physics, Rio de Janeiro (Brazil)

    1963-01-15

    An attractive feature common to all present theories of weak interactions is the assumption that they result from the coupling (either direct or via intermediate bosons) of vector-axial vector currents. The V-A coupling of leptonic currents and baryonic currents conserving strangeness successfully explains the characteristic properties of μ-decay, π-decay and μ-decay. On the other hand, the leptonic decay modes of K-mesons provide the best source of information on the nature of the currents carrying strangeness. From K{sub μ2} one learns that there must exist an axial-vector current carrying strangeness; further, if the leptonic weak interaction is universal, one can explain the absence of the K{sub e2} mode by excluding a pseudo-scalar interaction. From K{sub μ3} and K{sub e3} decay one can obtain information on scalar, vector and tensor coupling of strangeness non-conserving currents with leptons. In recent experiments on K{sub e3} decay a good fitting of the pion spectrum was obtained with a constant vector form factor. Although regarding this result as strong evidence of pure vector coupling, one might still argue that other possibilities, such as a mixture of vector and tensor couplings with energy-dependent form factors, are not excluded. I shall discuss here, in the first place, some properties of the transition amplitudes which are independent of the structure of form factors. They provide a test of universality and a criterion for unambiguous determination of the nature of the weak interaction involved in these processes. Secondly, I shall discuss the structure of form factors, by means of dispersion relations and introducing explicitly the effect of the K*-resonance in the Kπ-interaction.

  13. The APC/C E3 Ligase Complex Activator FZR1 Restricts BRAF Oncogenic Function.

    Science.gov (United States)

    Wan, Lixin; Chen, Ming; Cao, Juxiang; Dai, Xiangpeng; Yin, Qing; Zhang, Jinfang; Song, Su-Jung; Lu, Ying; Liu, Jing; Inuzuka, Hiroyuki; Katon, Jesse M; Berry, Kelsey; Fung, Jacqueline; Ng, Christopher; Liu, Pengda; Song, Min Sup; Xue, Lian; Bronson, Roderick T; Kirschner, Marc W; Cui, Rutao; Pandolfi, Pier Paolo; Wei, Wenyi

    2017-04-01

    BRAF drives tumorigenesis by coordinating the activation of the RAS/RAF/MEK/ERK oncogenic signaling cascade. However, upstream pathways governing BRAF kinase activity and protein stability remain undefined. Here, we report that in primary cells with active APC FZR1 , APC FZR1 earmarks BRAF for ubiquitination-mediated proteolysis, whereas in cancer cells with APC-free FZR1, FZR1 suppresses BRAF through disrupting BRAF dimerization. Moreover, we identified FZR1 as a direct target of ERK and CYCLIN D1/CDK4 kinases. Phosphorylation of FZR1 inhibits APC FZR1 , leading to elevation of a cohort of oncogenic APC FZR1 substrates to facilitate melanomagenesis. Importantly, CDK4 and/or BRAF/MEK inhibitors restore APC FZR1 E3 ligase activity, which might be critical for their clinical effects. Furthermore, FZR1 depletion cooperates with AKT hyperactivation to transform primary melanocytes, whereas genetic ablation of Fzr1 synergizes with Pten loss, leading to aberrant coactivation of BRAF/ERK and AKT signaling in mice. Our findings therefore reveal a reciprocal suppression mechanism between FZR1 and BRAF in controlling tumorigenesis. Significance: FZR1 inhibits BRAF oncogenic functions via both APC-dependent proteolysis and APC-independent disruption of BRAF dimers, whereas hyperactivated ERK and CDK4 reciprocally suppress APC FZR1 E3 ligase activity. Aberrancies in this newly defined signaling network might account for BRAF hyperactivation in human cancers, suggesting that targeting CYCLIN D1/CDK4, alone or in combination with BRAF/MEK inhibition, can be an effective anti-melanoma therapy. Cancer Discov; 7(4); 424-41. ©2017 AACR. See related commentary by Zhang and Bollag, p. 356 This article is highlighted in the In This Issue feature, p. 339 . ©2017 American Association for Cancer Research.

  14. E2-EPF UCP Possesses E3 Ubiquitin Ligase Activity via Its Cysteine 118 Residue.

    Science.gov (United States)

    Lim, Jung Hwa; Shin, Hee Won; Chung, Kyung-Sook; Kim, Nam-Soon; Kim, Ju Hee; Jung, Hong-Ryul; Im, Dong-Soo; Jung, Cho-Rok

    Here, we show that E2-EPF ubiquitin carrier protein (UCP) elongated E3-independent polyubiquitin chains on the lysine residues of von Hippel-Lindau protein (pVHL) and its own lysine residues both in vitro and in vivo. The initiation of the ubiquitin reaction depended on not only Lys11 linkage but also the Lys6, Lys48 and Lys63 residues of ubiquitin, which were involved in polyubiquitin chain formation on UCP itself. UCP self-association occurred through the UBC domain, which also contributed to the interaction with pVHL. The polyubiquitin chains appeared on the N-terminus of UCP in vivo, which indicated that the N-terminus of UCP contains target lysines for polyubiquitination. The Lys76 residue of UCP was the most critical site for auto-ubiquitination, whereas the polyubiquitin chain formation on pVHL occurred on all three of its lysines (Lys159, Lys171 and Lys196). A UCP mutant in which Cys118 was changed to alanine (UCPC118A) did not form a polyubiquitin chain but did strongly accumulate mono- and di-ubiquitin via auto-ubiquitination. Polyubiquitin chain formation required the coordination of Cys95 and Cys118 between two interacting molecules. The mechanism of the polyubiquitin chain reaction of UCP may involve the transfer of ubiquitin from Cys95 to Cys118 by trans-thiolation, with polyubiquitin chains forming at Cys118 by reversible thioester bonding. The polyubiquitin chains are then moved to the lysine residues of the substrate by irreversible isopeptide bonding. During the elongation of the ubiquitin chain, an active Cys118 residue is required in both parts of UCP, namely, the catalytic enzyme and the substrate. In conclusion, UCP possesses not only E2 ubiquitin conjugating enzyme activity but also E3 ubiquitin ligase activity, and Cys118 is critical for polyubiquitin chain formation.

  15. E2-EPF UCP Possesses E3 Ubiquitin Ligase Activity via Its Cysteine 118 Residue.

    Directory of Open Access Journals (Sweden)

    Jung Hwa Lim

    Full Text Available Here, we show that E2-EPF ubiquitin carrier protein (UCP elongated E3-independent polyubiquitin chains on the lysine residues of von Hippel-Lindau protein (pVHL and its own lysine residues both in vitro and in vivo. The initiation of the ubiquitin reaction depended on not only Lys11 linkage but also the Lys6, Lys48 and Lys63 residues of ubiquitin, which were involved in polyubiquitin chain formation on UCP itself. UCP self-association occurred through the UBC domain, which also contributed to the interaction with pVHL. The polyubiquitin chains appeared on the N-terminus of UCP in vivo, which indicated that the N-terminus of UCP contains target lysines for polyubiquitination. The Lys76 residue of UCP was the most critical site for auto-ubiquitination, whereas the polyubiquitin chain formation on pVHL occurred on all three of its lysines (Lys159, Lys171 and Lys196. A UCP mutant in which Cys118 was changed to alanine (UCPC118A did not form a polyubiquitin chain but did strongly accumulate mono- and di-ubiquitin via auto-ubiquitination. Polyubiquitin chain formation required the coordination of Cys95 and Cys118 between two interacting molecules. The mechanism of the polyubiquitin chain reaction of UCP may involve the transfer of ubiquitin from Cys95 to Cys118 by trans-thiolation, with polyubiquitin chains forming at Cys118 by reversible thioester bonding. The polyubiquitin chains are then moved to the lysine residues of the substrate by irreversible isopeptide bonding. During the elongation of the ubiquitin chain, an active Cys118 residue is required in both parts of UCP, namely, the catalytic enzyme and the substrate. In conclusion, UCP possesses not only E2 ubiquitin conjugating enzyme activity but also E3 ubiquitin ligase activity, and Cys118 is critical for polyubiquitin chain formation.

  16. Phosphorylation by PINK1 releases the UBL domain and initializes the conformational opening of the E3 ubiquitin ligase Parkin.

    Directory of Open Access Journals (Sweden)

    Thomas R Caulfield

    2014-11-01

    Full Text Available Loss-of-function mutations in PINK1 or PARKIN are the most common causes of autosomal recessive Parkinson's disease. Both gene products, the Ser/Thr kinase PINK1 and the E3 Ubiquitin ligase Parkin, functionally cooperate in a mitochondrial quality control pathway. Upon stress, PINK1 activates Parkin and enables its translocation to and ubiquitination of damaged mitochondria to facilitate their clearance from the cell. Though PINK1-dependent phosphorylation of Ser65 is an important initial step, the molecular mechanisms underlying the activation of Parkin's enzymatic functions remain unclear. Using molecular modeling, we generated a complete structural model of human Parkin at all atom resolution. At steady state, the Ub ligase is maintained inactive in a closed, auto-inhibited conformation that results from intra-molecular interactions. Evidently, Parkin has to undergo major structural rearrangements in order to unleash its catalytic activity. As a spark, we have modeled PINK1-dependent Ser65 phosphorylation in silico and provide the first molecular dynamics simulation of Parkin conformations along a sequential unfolding pathway that could release its intertwined domains and enable its catalytic activity. We combined free (unbiased molecular dynamics simulation, Monte Carlo algorithms, and minimal-biasing methods with cell-based high content imaging and biochemical assays. Phosphorylation of Ser65 results in widening of a newly defined cleft and dissociation of the regulatory N-terminal UBL domain. This motion propagates through further opening conformations that allow binding of an Ub-loaded E2 co-enzyme. Subsequent spatial reorientation of the catalytic centers of both enzymes might facilitate the transfer of the Ub moiety to charge Parkin. Our structure-function study provides the basis to elucidate regulatory mechanisms and activity of the neuroprotective Parkin. This may open up new avenues for the development of small molecule Parkin

  17. Inhibition of xanthine oxidase by allopurinol prevents skeletal muscle atrophy: role of p38 MAPKinase and E3 ubiquitin ligases.

    Directory of Open Access Journals (Sweden)

    Frederic Derbre

    Full Text Available Alterations in muscle play an important role in common diseases and conditions. Reactive oxygen species (ROS are generated during hindlimb unloading due, at least in part, to the activation of xanthine oxidase (XO. The major aim of this study was to determine the mechanism by which XO activation causes unloading-induced muscle atrophy in rats, and its possible prevention by allopurinol, a well-known inhibitor of this enzyme. For this purpose we studied one of the main redox sensitive signalling cascades involved in skeletal muscle atrophy i.e. p38 MAPKinase, and the expression of two well known muscle specific E3 ubiquitin ligases involved in proteolysis, the Muscle atrophy F-Box (MAFbx; also known as atrogin-1 and Muscle RING (Really Interesting New Gene Finger-1 (MuRF-1. We found that hindlimb unloading induced a significant increase in XO activity and in the protein expression of the antioxidant enzymes CuZnSOD and Catalase in skeletal muscle. The most relevant new fact reported in this paper is that inhibition of XO with allopurinol, a drug widely used in clinical practice, prevents soleus muscle atrophy by ~20% after hindlimb unloading. This was associated with the inhibition of the p38 MAPK-MAFbx pathway. Our data suggest that XO was involved in the loss of muscle mass via the activation of the p38MAPK-MAFbx pathway in unloaded muscle atrophy. Thus, allopurinol may have clinical benefits to combat skeletal muscle atrophy in bedridden, astronauts, sarcopenic, and cachexic patients.

  18. Arabidopsis C3HC4-RING finger E3 ubiquitin ligase AtAIRP4 positively regulates stress-responsive abscisic acid signaling.

    Science.gov (United States)

    Yang, Liang; Liu, Qiaohong; Liu, Zhibin; Yang, Hao; Wang, Jianmei; Li, Xufeng; Yang, Yi

    2016-01-01

    Degradation of proteins via the ubiquitin system is an important step in many stress signaling pathways in plants. E3 ligases recognize ligand proteins and dictate the high specificity of protein degradation, and thus, play a pivotal role in ubiquitination. Here, we identified a gene, named Arabidopsis thaliana abscisic acid (ABA)-insensitive RING protein 4 (AtAIRP4), which is induced by ABA and other stress treatments. AtAIRP4 encodes a cellular protein with a C3HC4-RING finger domain in its C-terminal side, which has in vitro E3 ligase activity. Loss of AtAIRP4 leads to a decrease in sensitivity of root elongation and stomatal closure to ABA, whereas overexpression of this gene in the T-DNA insertion mutant atairp4 effectively recovered the ABA-associated phenotypes. AtAIRP4 overexpression plants were hypersensitive to salt and osmotic stresses during seed germination, and showed drought avoidance compared with the wild-type and atairp4 mutant plants. In addition, the expression levels of ABA- and drought-induced marker genes in AtAIRP4 overexpression plants were markedly higher than those in the wild-type and atairp4 mutant plants. Hence, these results indicate that AtAIRP4 may act as a positive regulator of ABA-mediated drought avoidance and a negative regulator of salt tolerance in Arabidopsis. © 2015 The Authors. Journal of Integrative Plant Biology published by Wiley Publishing Asia Pty Ltd on behalf of Institute of Botany, Chinese Academy of Sciences.

  19. The Oncogenic Role of WWP1 E3 Ubiquitin Ligase in Prostate Cancer Development

    Science.gov (United States)

    2011-05-01

    screemng method. Drosophila S2 cetls were trall$lenUy transfected with the proteasome UFO substrate UbG76VGFP to Identify transcription factors that...specialanterest were three genes, since knock down of these showed the strongest stabilization of the UFO substrate. These were FOXO, a1c and maf-S

  20. 10 CFR 830 Major Modification Determination for the ATR Diesel Bus (E-3) and Switchgear Replacement

    International Nuclear Information System (INIS)

    Duckwitz, Noel

    2011-01-01

    Near term replacement of aging and obsolescent original ATR equipment has become important to ensure ATR capability in support of NE's long term national missions. To that end, a mission needs statement has been prepared for a non-major system acquisition which is comprised of three interdependent subprojects. The first project, subject of this determination, will replace the existent diesel-electrical bus (E-3) and associated switchgear. More specifically, INL proposes transitioning ATR to 100% commercial power with appropriate emergency backup to include: (1) Provide commercial power as the normal source of power to the ATR loads currently supplied by diesel-electric power. (2) Provide backup power to the critical ATR loads in the event of a loss of commercial power. (3) Replace obsolescent critical ATR power distribution equipment, e.g., switchgear, transformers, motor control centers, distribution panels. Completion of this and two other age-related projects (primary coolant pump and motor replacement and emergency firewater injection system replacement) will resolve major age related operational issues plus make a significant contribution in sustaining the ATR safety and reliability profile. The major modification criteria evaluation of the project pre-conceptual design identified several issues make the project a major modification: (1) Evaluation Criteria No.2 (Footprint change). The addition of a new PC-4 structure to the ATR Facility to house safety-related SSCs requires careful attention to maintaining adherence to applicable engineering and nuclear safety design criteria (e.g., structural qualification, fire suppression) to ensure no adverse impacts to the safety-related functions of the housed equipment. (2) Evaluation Criteria No.3 (Change of existing process). The change to the strategy for providing continuous reliable power to the safety-related emergency coolant pumps requires careful attention and analysis to ensure it meets a project primary

  1. Diagnostic Value of Leukocyte Esterase Test Strip Reagents for Rapid Clinical Diagnosis of Spontaneous Bacterial Peritonitis in Patients Admitted to Hospital Emergency Departments in Iran.

    Science.gov (United States)

    Hashemian, Amir Masoud; Ahmadi, Koorosh; Zamani Moghaddam, Hamid; Zakeri, Hosein; Davoodi Navakh, Seyed Akbar; Sharifi, Mohammad Davood; Bahrami, Abdollah

    2015-10-01

    Spontaneous bacterial peritonitis (SBP) is a common and important clinical problem and is life-threatening in decompensated liver disease. Ascites fluid test by leukocyte esterase test strip has been recently proposed as an effective and rapid method to diagnose SBP in patients with cirrhosis. This study aimed to evaluate sensitivity and specificity of leukocyte esterase test strip in the diagnosis of SBP. The population of this research was all patients with cirrhosis and ascites admitted to the emergency room at Imam Reza (AS) hospital, Mashhad. A written consent was taken for inclusion in the study. 50 mL ascites sample was taken from all patients for use in a urine test strip (LER) (Urine Test Strips Convergys®Urine Matrix 11). The patient's ascites samples were evaluated for cell counting. Positive dipstick test for LER in this study considered as grade 3 +. The values of WBC > 500 cell/mm(3) or PMN > 250 cell/mm(3) considered as positive result of the gold standard method for the diagnosis of SBP. In this study, 100 patients with ascites due to cirrhosis, with an average age of 38.9 ± 6.54 years were evaluated. Twenty cases had positive results, of whom 17 cases were also detected based on the standard diagnostic criteria and other three cases were healthy individuals. Thus, sensitivity, specificity, positive and negative predictive values, and accuracy of the method were 95%, 96.3%, 85%, 97.5% and 95%, respectively. The use of leukocyte esterase urine dipstick test can be a quick and easy method in early diagnosis of SBP to start the treatment until preparation of SBP-cell count results.

  2. Geranyl acetate esterase controls and regulates the level of geraniol in lemongrass (Cymbopogon flexuosus Nees ex Steud.) mutant cv. GRL-1 leaves.

    Science.gov (United States)

    Ganjewala, Deepak; Luthra, Rajesh

    2009-01-01

    Essential oil isolated from lemongrass (Cymbopogon flexuosus) mutant cv. GRL-1 leaves is mainly composed of geraniol (G) and geranyl acetate (GA). The proportion of G and GA markedly fluctuates during leaf development. The proportions of GA and G in the essential oil recorded at day 10 after leaf emergence were approximately 59% and approximately 33% respectively. However, the level of GA went down from approximately 59 to approximately 3% whereas the level of G rose from approximately 33 to approximately 91% during the leaf growth period from day 10 to day 50. However, the decline in the level of GA was most pronounced in the early (day 10 to day 30) stage of leaf growth. The trend of changes in the proportion of GA and G has clearly indicated the role of an esterase that must be involved in the conversion of GA to G during leaf development. We isolated an esterase from leaves of different ages that converts GA into G and has been given the name geranyl acetate esterase (GAE). The GAE activity markedly varied during the leaf development cycle; it was closely correlated with the monoterpene (GA and G) composition throughout leaf development. GAE appeared as several isoenzymes but only three (GAE-I, GAE-II, and GAE-III) of them had significant GA cleaving activity. The GAE isoenzymes pattern was greatly influenced by the leaf developmental stages and so their GA cleaving activities. Like the GAE activity, GAE isoenzyme patterns were also found to be consistent with the monoterpene (GA and G) composition. GAE had an optimum pH at 8.5 and temperature at 30 degrees C. Besides GAE, a compound with phosphatase activity capable of hydrolyzing geranyl diphosphate (GPP) to produce geraniol has also been isolated.

  3. The Alpha-Defensin Immunoassay and Leukocyte Esterase Colorimetric Strip Test for the Diagnosis of Periprosthetic Infection: A Systematic Review and Meta-Analysis.

    Science.gov (United States)

    Wyatt, M C; Beswick, A D; Kunutsor, S K; Wilson, M J; Whitehouse, M R; Blom, A W

    2016-06-15

    Synovial biomarkers have recently been adopted as diagnostic tools for periprosthetic joint infection (PJI), but their utility is uncertain. The purpose of this systematic review and meta-analysis was to synthesize the evidence on the accuracy of the alpha-defensin immunoassay and leukocyte esterase colorimetric strip test for the diagnosis of PJI compared with the Musculoskeletal Infection Society diagnostic criteria. We performed a systematic review to identify diagnostic technique studies evaluating the accuracy of alpha-defensin or leukocyte esterase in the diagnosis of PJI. MEDLINE and Embase on Ovid, ACM, ADS, arXiv, CERN DS (Conseil Européen pour la Recherche Nucléaire Document Server), CrossRef DOI (Digital Object Identifier), DBLP (Digital Bibliography & Library Project), Espacenet, Google Scholar, Gutenberg, HighWire, IEEE Xplore (Institute of Electrical and Electronics Engineers digital library), INSPIRE, JSTOR (Journal Storage), OAlster (Open Archives Initiative Protocol for Metadata Harvesting), Open Content, Pubget, PubMed, and Web of Science were searched for appropriate studies indexed from inception until May 30, 2015, along with unpublished or gray literature. The classification of studies and data extraction were performed independently by 2 reviewers. Data extraction permitted meta-analysis of sensitivity and specificity with construction of receiver operating characteristic curves for each test. We included 11 eligible studies. The pooled diagnostic sensitivity and specificity of alpha-defensin (6 studies) for PJI were 1.00 (95% confidence interval [CI], 0.82 to 1.00) and 0.96 (95% CI, 0.89 to 0.99), respectively. The area under the curve (AUC) for alpha-defensin and PJI was 0.99 (95% CI, 0.98 to 1.00). The pooled diagnostic sensitivity and specificity of leukocyte esterase (5 studies) for PJI were 0.81 (95% CI, 0.49 to 0.95) and 0.97 (95% CI, 0.82 to 0.99), respectively. The AUC for leukocyte esterase and PJI was 0.97 (95% CI, 0.95 to 0

  4. Changes in pectin methyl esterase activity with different packaging materials and stages of fruit harvesting during cold storage of pear cv. Punjab beauty

    OpenAIRE

    Kaur, Kirandeep; Dhillon, W. S.; Mahajan, B. V. C.

    2012-01-01

    Pear cv. Punjab Beauty has become quite popular in Punjab. Excessive softening during cold storage leading to low shelf life is the major factor limiting its wider adoption. Studies were, therefore, conducted to determine the firmness and pectin methyl esterase (PME) activity at 4 harvest dates (2nd, 3rd and 4th week of July, and 1st week of August). Various packaging materials i.e. corrugated fiber board boxes and crates with high and low density polyethylene liners, corrugated fiber board b...

  5. Kinetics of the inhibitory interaction of organophosphorus neuropathy inducers and non-inducers in soluble esterases in the avian nervous system

    Energy Technology Data Exchange (ETDEWEB)

    Mangas, Iris; Vilanova, Eugenio; Estevez, Jorge, E-mail: jorge.estevez@umh.es

    2011-11-15

    Some published studies suggest that low level exposure to organophosphorus esters (OPs) may cause neurological and neurobehavioral effects at long term exposure. These effects cannot be explained by action on known targets. In this work, the interactions (inhibition, spontaneous reactivation and 'ongoing inhibition') of two model OPs (paraoxon, non neuropathy-inducer, and mipafox, neuropathy-inducer) with the chicken brain soluble esterases were evaluated. The best-fitting kinetic model with both inhibitors was compatible with three enzymatic components. The amplitudes (proportions) of the components detected with mipafox were similar to those obtained with paraoxon. These observations confirm the consistency of the results and the model applied and may be considered an external validation. The most sensitive component (E{alpha}) for paraoxon (11-23% of activity, I{sub 50} (30 min) = 9-11 nM) is also the most sensitive for mipafox (I{sub 50} (30 min) = 4 nM). This component is spontaneously reactivated after inhibition with paraoxon. The second sensitive component to paraoxon (E{beta}, 71-84% of activity; I{sub 50} (30 min) = 1216 nM) is practically resistant to mipafox. The third component (E{gamma}, 5-8% of activity) is paraoxon resistant and has I{sub 50} (30 min) of 3.4 {mu}M with mipafox, similar to NTE (neuropathy target esterase). The role of these esterases remains unknown. Their high sensitivity suggests that they may either play a role in toxicity in low-level long-term exposure of organophosphate compounds or have a protective effect related with the spontaneous reactivation. They will have to be considered in further metabolic and toxicological studies. -- Research Highlights: Black-Right-Pointing-Pointer Paraoxon and mipafox interactions have been evaluated with chicken soluble brain esterases. Black-Right-Pointing-Pointer The paraoxon inhibition was analyzed considering the simultaneous spontaneous reactivation. Black

  6. Kinetics of the inhibitory interaction of organophosphorus neuropathy inducers and non-inducers in soluble esterases in the avian nervous system

    International Nuclear Information System (INIS)

    Mangas, Iris; Vilanova, Eugenio; Estévez, Jorge

    2011-01-01

    Some published studies suggest that low level exposure to organophosphorus esters (OPs) may cause neurological and neurobehavioral effects at long term exposure. These effects cannot be explained by action on known targets. In this work, the interactions (inhibition, spontaneous reactivation and “ongoing inhibition”) of two model OPs (paraoxon, non neuropathy-inducer, and mipafox, neuropathy-inducer) with the chicken brain soluble esterases were evaluated. The best-fitting kinetic model with both inhibitors was compatible with three enzymatic components. The amplitudes (proportions) of the components detected with mipafox were similar to those obtained with paraoxon. These observations confirm the consistency of the results and the model applied and may be considered an external validation. The most sensitive component (Eα) for paraoxon (11–23% of activity, I 50 (30 min) = 9–11 nM) is also the most sensitive for mipafox (I 50 (30 min) = 4 nM). This component is spontaneously reactivated after inhibition with paraoxon. The second sensitive component to paraoxon (Eβ, 71–84% of activity; I 50 (30 min) = 1216 nM) is practically resistant to mipafox. The third component (Eγ, 5–8% of activity) is paraoxon resistant and has I 50 (30 min) of 3.4 μM with mipafox, similar to NTE (neuropathy target esterase). The role of these esterases remains unknown. Their high sensitivity suggests that they may either play a role in toxicity in low-level long-term exposure of organophosphate compounds or have a protective effect related with the spontaneous reactivation. They will have to be considered in further metabolic and toxicological studies. -- Research Highlights: ► Paraoxon and mipafox interactions have been evaluated with chicken soluble brain esterases. ► The paraoxon inhibition was analyzed considering the simultaneous spontaneous reactivation. ► The best-fitting kinetic models were compatible with a three enzymatic components. ► The amplitudes of the

  7. Sustainable energy development in Austria until 2020: Insights from applying the integrated model 'e3.at'

    International Nuclear Information System (INIS)

    Stocker, Andrea; Grossmann, Anett; Madlener, Reinhard; Wolter, Marc Ingo

    2011-01-01

    This paper reports on the Austrian research project 'Renewable energy in Austria: Modeling possible development trends until 2020'. The project investigated possible economic and ecological effects of a substantially increased use of renewable energy sources in Austria. Together with stakeholders and experts, three different scenarios were defined, specifying possible development trends for renewable energy in Austria. The scenarios were simulated for the period 2006-2020, using the integrated environment-energy-economy model 'e3.at'. The modeling results indicate that increasing the share of renewable energy sources in total energy use is an important but insufficient step towards achieving a sustainable energy system in Austria. A substantial increase in energy efficiency and a reduction of residential energy consumption also form important cornerstones of a sustainable energy policy. - Highlights: → Together with stakeholders three renewable energy scenarios for Austria were defined. → The scenarios were simulated using an integrated environment-energy-economy model. → Increasing the share of renewables in total energy use is important but insufficient. → Efficiency and a cut of energy use are also essential for a sustainable energy system.

  8. Correlated compositional and mineralogical investigations at the Chang′e-3 landing site

    Science.gov (United States)

    Ling, Zongcheng; Jolliff, Bradley L.; Wang, Alian; Li, Chunlai; Liu, Jianzhong; Zhang, Jiang; Li, Bo; Sun, Lingzhi; Chen, Jian; Xiao, Long; Liu, Jianjun; Ren, Xin; Peng, Wenxi; Wang, Huanyu; Cui, Xingzhu; He, Zhiping; Wang, Jianyu

    2015-01-01

    The chemical compositions of relatively young mare lava flows have implications for the late volcanism on the Moon. Here we report the composition of soil along the rim of a 450-m diameter fresh crater at the Chang′e-3 (CE-3) landing site, investigated by the Yutu rover with in situ APXS (Active Particle-induced X-ray Spectrometer) and VNIS (Visible and Near-infrared Imaging Spectrometer) measurements. Results indicate that this region's composition differs from other mare sample-return sites and is a new type of mare basalt not previously sampled, but consistent with remote sensing. The CE-3 regolith derived from olivine-normative basaltic rocks with high FeO/(FeO+MgO). Deconvolution of the VNIS data indicates abundant high-Ca ferropyroxene (augite and pigeonite) plus Fe-rich olivine. We infer from the regolith composition that the basaltic source rocks formed during late-stage magma-ocean differentiation when dense ferropyroxene-ilmenite cumulates sank and mixed with deeper, relatively ferroan olivine and orthopyroxene in a hybridized mantle source. PMID:26694712

  9. Preface: The Chang'e-3 lander and rover mission to the Moon

    International Nuclear Information System (INIS)

    Ip Wing-Huen; Yan Jun; Li Chun-Lai; Ouyang Zi-Yuan

    2014-01-01

    The Chang'e-3 (CE-3) lander and rover mission to the Moon was an intermediate step in China's lunar exploration program, which will be followed by a sample return mission. The lander was equipped with a number of remote-sensing instruments including a pair of cameras (Landing Camera and Terrain Camera) for recording the landing process and surveying terrain, an extreme ultraviolet camera for monitoring activities in the Earth's plasmasphere, and a first-ever Moon-based ultraviolet telescope for astronomical observations. The Yutu rover successfully carried out close-up observations with the Panoramic Camera, mineralogical investigations with the VIS-NIR Imaging Spectrometer, study of elemental abundances with the Active Particle-induced X-ray Spectrometer, and pioneering measurements of the lunar subsurface with Lunar Penetrating Radar. This special issue provides a collection of key information on the instrumental designs, calibration methods and data processing procedures used by these experiments with a perspective of facilitating further analyses of scientific data from CE-3 in preparation for future missions

  10. Correlated compositional and mineralogical investigations at the Chang'e-3 landing site.

    Science.gov (United States)

    Ling, Zongcheng; Jolliff, Bradley L; Wang, Alian; Li, Chunlai; Liu, Jianzhong; Zhang, Jiang; Li, Bo; Sun, Lingzhi; Chen, Jian; Xiao, Long; Liu, Jianjun; Ren, Xin; Peng, Wenxi; Wang, Huanyu; Cui, Xingzhu; He, Zhiping; Wang, Jianyu

    2015-12-22

    The chemical compositions of relatively young mare lava flows have implications for the late volcanism on the Moon. Here we report the composition of soil along the rim of a 450-m diameter fresh crater at the Chang'e-3 (CE-3) landing site, investigated by the Yutu rover with in situ APXS (Active Particle-induced X-ray Spectrometer) and VNIS (Visible and Near-infrared Imaging Spectrometer) measurements. Results indicate that this region's composition differs from other mare sample-return sites and is a new type of mare basalt not previously sampled, but consistent with remote sensing. The CE-3 regolith derived from olivine-normative basaltic rocks with high FeO/(FeO+MgO). Deconvolution of the VNIS data indicates abundant high-Ca ferropyroxene (augite and pigeonite) plus Fe-rich olivine. We infer from the regolith composition that the basaltic source rocks formed during late-stage magma-ocean differentiation when dense ferropyroxene-ilmenite cumulates sank and mixed with deeper, relatively ferroan olivine and orthopyroxene in a hybridized mantle source.

  11. SUMO E3 ligase Mms21 prevents spontaneous DNA damage induced genome rearrangements.

    Directory of Open Access Journals (Sweden)

    Jason Liang

    2018-03-01

    Full Text Available Mms21, a subunit of the Smc5/6 complex, possesses an E3 ligase activity for the Small Ubiquitin-like MOdifier (SUMO. Here we show that the mms21-CH mutation, which inactivates Mms21 ligase activity, causes increased accumulation of gross chromosomal rearrangements (GCRs selected in the dGCR assay. These dGCRs are formed by non-allelic homologous recombination between divergent DNA sequences mediated by Rad52-, Rrm3- and Pol32-dependent break-induced replication. Combining mms21-CH with sgs1Δ caused a synergistic increase in GCRs rates, indicating the distinct roles of Mms21 and Sgs1 in suppressing GCRs. The mms21-CH mutation also caused increased rates of accumulating uGCRs mediated by breakpoints in unique sequences as revealed by whole genome sequencing. Consistent with the accumulation of endogenous DNA lesions, mms21-CH mutants accumulate increased levels of spontaneous Rad52 and Ddc2 foci and had a hyper-activated DNA damage checkpoint. Together, these findings support that Mms21 prevents the accumulation of spontaneous DNA lesions that cause diverse GCRs.

  12. Preface: The Chang'e-3 lander and rover mission to the Moon

    Science.gov (United States)

    Ip, Wing-Huen; Yan, Jun; Li, Chun-Lai; Ouyang, Zi-Yuan

    2014-12-01

    The Chang'e-3 (CE-3) lander and rover mission to the Moon was an intermediate step in China's lunar exploration program, which will be followed by a sample return mission. The lander was equipped with a number of remote-sensing instruments including a pair of cameras (Landing Camera and Terrain Camera) for recording the landing process and surveying terrain, an extreme ultraviolet camera for monitoring activities in the Earth's plasmasphere, and a first-ever Moon-based ultraviolet telescope for astronomical observations. The Yutu rover successfully carried out close-up observations with the Panoramic Camera, mineralogical investigations with the VIS-NIR Imaging Spectrometer, study of elemental abundances with the Active Particle-induced X-ray Spectrometer, and pioneering measurements of the lunar subsurface with Lunar Penetrating Radar. This special issue provides a collection of key information on the instrumental designs, calibration methods and data processing procedures used by these experiments with a perspective of facilitating further analyses of scientific data from CE-3 in preparation for future missions.

  13. The Processing of Lunar Penetrating Radar Channel-2B Data from Chang'E-3

    Science.gov (United States)

    Zhu, P.; Zhao, N.; Yang, K.; Yuan, Y.; Guo, S.

    2014-12-01

    The Lunar Penetrating Radar (LPR) carried by Chang'E-3 has imaged the shallow subsurface of the landing site at the northern Mare Imbrium. The antenna B of the Channel-2 onboard the LPR (LPR Channel-2B) has collected more than 20000 traces of raw data. The raw LPR data could not be directly used for geological interpretation because of the operation mode of the LPR, noise and fast attenuation of radar wave. This study focuses data preprocessing and processing methods to obtain higher quality data. A section of usable LPR data of over 2000 traces is gained after the preprocessing of selecting, splicing, removing delay time and fine-correcting to raw data, but only a few shallow geological structures are visible. To further improve the resolution and the signal-to-noise ratio of the LPR data, we have processed the LPR data including amplitude compensation, filtering, and deconvolution processes based on electromagnetic wave theory. The processing results reveal that (1) the spherical spreading compensation and auto gain control enhance the amplitude of reflection echoes from deeper strata and make the geological structures more obvious, (2) the spiking deconvolution applied to narrow reflection events down makes it possible to identify thin layers with 30% improved resolution, and (3) the band-pass filtering removes the multiple reflections and, consequently, improves the signal-to-noise ratio of LPR data. The processing results will lay the foundation for the subsequent geological interpretation and physical property inversion of lunar materials.

  14. Deviations between experimental and theoretical results in Ar (e, 3e) double ionization

    International Nuclear Information System (INIS)

    Jia, C C; Lahmam-Bennani, A; Cappello, C Dal; Duguet, A; Avaldi, L

    2003-01-01

    The coplanar (e, 3e) relative cross sections for double ionization of argon have been measured at an electron impact energy of E 0 = 561.4 eV and under equal energy sharing among the two 'ejected' electrons, E b = E c = 9 eV. The scattering angle is fixed to θ a = 1.5 degrees, corresponding to a momentum transfer K=0.4 au to the target. The experimental results have been compared with calculations in the first Born approximation, which include only first-order processes in the target-projectile interaction. The comparison shows severe deviations between the experimental and theoretical results. These deviations are much larger than the ones previously observed in helium under comparable kinematics. To fill this gap between theory and experiment, a decisive improvement in the theory is needed. This can be achieved by improving the first-order calculations and by including higher-order processes such as the two-step mechanism, or even new mechanisms, for instance the simultaneous ejection of the pair of target electrons. (letter to the editor)

  15. The Impact of Parametric Uncertainties on Biogeochemistry in the E3SM Land Model

    Science.gov (United States)

    Ricciuto, Daniel; Sargsyan, Khachik; Thornton, Peter

    2018-02-01

    We conduct a global sensitivity analysis (GSA) of the Energy Exascale Earth System Model (E3SM), land model (ELM) to calculate the sensitivity of five key carbon cycle outputs to 68 model parameters. This GSA is conducted by first constructing a Polynomial Chaos (PC) surrogate via new Weighted Iterative Bayesian Compressive Sensing (WIBCS) algorithm for adaptive basis growth leading to a sparse, high-dimensional PC surrogate with 3,000 model evaluations. The PC surrogate allows efficient extraction of GSA information leading to further dimensionality reduction. The GSA is performed at 96 FLUXNET sites covering multiple plant functional types (PFTs) and climate conditions. About 20 of the model parameters are identified as sensitive with the rest being relatively insensitive across all outputs and PFTs. These sensitivities are dependent on PFT, and are relatively consistent among sites within the same PFT. The five model outputs have a majority of their highly sensitive parameters in common. A common subset of sensitive parameters is also shared among PFTs, but some parameters are specific to certain types (e.g., deciduous phenology). The relative importance of these parameters shifts significantly among PFTs and with climatic variables such as mean annual temperature.

  16. An assessment of Japanese carbon tax reform using the E3MG econometric model.

    Science.gov (United States)

    Lee, Soocheol; Pollitt, Hector; Ueta, Kazuhiro

    2012-01-01

    This paper analyses the potential economic and environmental effects of carbon taxation in Japan using the E3MG model, a global macroeconometric model constructed by the University of Cambridge and Cambridge Econometrics. The paper approaches the issues by considering first the impacts of the carbon tax in Japan introduced in 2012 and then the measures necessary to reduce Japan's emissions in line with its Copenhagen pledge of -25% compared to 1990 levels. The results from the model suggest that FY2012 Tax Reform has only a small impact on emission levels and no significant impact on GDP and employment. The potential costs of reducing emissions to meet the 25% reduction target for 2020 are quite modest, but noticeable. GDP falls by around 1.2% compared to the baseline and employment by 0.4% compared to the baseline. But this could be offset, with some potential economic benefits, if revenues are recycled efficiently. This paper considers two revenue recycling scenarios. The most positive outcome is if revenues are used both to reduce income tax rates and to increase investment in energy efficiency. This paper shows there could be double dividend effects, if Carbon Tax Reform is properly designed.

  17. An Assessment of Japanese Carbon Tax Reform Using the E3MG Econometric Model

    Directory of Open Access Journals (Sweden)

    Soocheol Lee

    2012-01-01

    Full Text Available This paper analyses the potential economic and environmental effects of carbon taxation in Japan using the E3MG model, a global macroeconometric model constructed by the University of Cambridge and Cambridge Econometrics. The paper approaches the issues by considering first the impacts of the carbon tax in Japan introduced in 2012 and then the measures necessary to reduce Japan’s emissions in line with its Copenhagen pledge of −25% compared to 1990 levels. The results from the model suggest that FY2012 Tax Reform has only a small impact on emission levels and no significant impact on GDP and employment. The potential costs of reducing emissions to meet the 25% reduction target for 2020 are quite modest, but noticeable. GDP falls by around 1.2% compared to the baseline and employment by 0.4% compared to the baseline. But this could be offset, with some potential economic benefits, if revenues are recycled efficiently. This paper considers two revenue recycling scenarios. The most positive outcome is if revenues are used both to reduce income tax rates and to increase investment in energy efficiency. This paper shows there could be double dividend effects, if Carbon Tax Reform is properly designed.

  18. An Assessment of Japanese Carbon Tax Reform Using the E3MG Econometric Model

    Science.gov (United States)

    Lee, Soocheol; Pollitt, Hector; Ueta, Kazuhiro

    2012-01-01

    This paper analyses the potential economic and environmental effects of carbon taxation in Japan using the E3MG model, a global macroeconometric model constructed by the University of Cambridge and Cambridge Econometrics. The paper approaches the issues by considering first the impacts of the carbon tax in Japan introduced in 2012 and then the measures necessary to reduce Japan's emissions in line with its Copenhagen pledge of −25% compared to 1990 levels. The results from the model suggest that FY2012 Tax Reform has only a small impact on emission levels and no significant impact on GDP and employment. The potential costs of reducing emissions to meet the 25% reduction target for 2020 are quite modest, but noticeable. GDP falls by around 1.2% compared to the baseline and employment by 0.4% compared to the baseline. But this could be offset, with some potential economic benefits, if revenues are recycled efficiently. This paper considers two revenue recycling scenarios. The most positive outcome is if revenues are used both to reduce income tax rates and to increase investment in energy efficiency. This paper shows there could be double dividend effects, if Carbon Tax Reform is properly designed. PMID:23365531

  19. Reaction (γ,2e) and (e,3e) as probe of electron correlation in atoms

    International Nuclear Information System (INIS)

    Amusia, M.Y.

    1995-01-01

    Cross sections of the (γ,2e) and (e,3e) reactions contain information about the two vacancy-energy spectrum and electron-pair correlations in initial and final states of the target atom. Physical pictures of these processes are presented for two- and many-electron atoms. The simplest mechanisms are discussed, demonstrating some features which await experimental confirmation. Attention is given to high photon energy and the relativistic energy region of these reactions. The energy distribution of outgoing relativistic electrons is qualitatively different from the nonrelativistic case. The origin and types of corrections to the simplest mechanisms, and possible means of their detection, are discussed. In addition, the role of different resonances: shape, giant, autoionizational, and Feshbach-type are considered. Results of calculations are compared with experimental data, mainly on double photoionization cross sections. Different possible objects as targets for the reactions are considered, including negative ions, excited atoms, molecules, and clusters. The modification of these reactions due to photon emission is discussed. The future of the domain is outlined

  20. Reactions (γ,2e) and (e,3e) as probes of electronic correlations in atoms

    International Nuclear Information System (INIS)

    Amusia, M.Ya.

    1993-01-01

    Cross sections of the (γ,2e) and (e,3e) reactions carry information on two vacancy energy spectrum and on electron pair correlations in initial and final states of the target atom. Physical pictures of these processes are presented for two- and many-electron atoms. Simplest mechanisms of them are discussed, demonstrating some features which are waiting for experimental confirmation. Attention is given to high photon energy and even to relativistic energy region of these reactions. The energy distribution of outgoing relativistic electrons is qualitatively different from what it is for the nonrelativistic case. Origin and types of corrections to the simplest mechanisms and possible means of their detection are discussed. Role of different resonances: shape, giant, autoionizational, and Feschbach-type are considered. Results of calculations are compared with experimental data, mainly on double photoionization cross sections. Different possible objects as targets for the reactions are mentioned, including negative ions, excited atoms, molecules and clusters. Modification of the type of these reactions due to rather probable emission of the photon is discussed. Future of the domain is outlined. (orig.)